Oxadiazole derivatives

ABSTRACT

The invention relates to oxadiazole compounds of formula I. The compounds are useful e.g. in the treatment of autoimmune disorders, such as multiple sclerosis.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is the U.S. national stage application of InternationalPatent Application No. PCT/EP2010/054103, filed Mar. 29, 2010, whichclaims the benefit of U.S. Provisional Patent Application No.61/169,773, filed Apr. 16, 2009.

The present invention relates to oxadiazoles, their use as medicamentsand their use for treating multiple sclerosis and other diseases.

In particular, the invention relates to compounds of formula (I):

Wherein

-   -   R¹, R² independently from one another denote H, Hal, CF₃, OCF₃,        CN, or NO₂, OH,    -   S is OR³ or COOR³,    -   Q denotes (CH₂)_(m)X(CH₂)_(m*),    -   X is —NA-,    -   W denotes CH,    -   R^(a) is

-   -   wherein R^(s) denotes Hal, A, OR³, N(R³)₂, NO₂, CN, COOR³, CF₃,        OCF₃, CON(R³)₂, NR³COA, NR³CON(R³)₂, NR³SO₂A, COR³, SO₂N(R³)₂,        SOA or SO₂A, and G₁, G₂ independently from one another denote H,        Hal, or CH₃, preferably H,    -   R^(b) is A, CF₃, CH₃, Hal, OCF₃, CN, NO₂, N(R³)₂, CH₂(NR₃)₂,    -   A denotes a linear or branched alkyl having 1 to 6 C-atoms,        preferably 1 to 3 carbon atoms, wherein one H-atom may be        replaced by Hal, OH, COOR³, CN, N(R³)₂ and wherein one CH₂—        group may be replaced by O, —NR³—, —NR³CO—, —CO— or S and/or by        —CH═CH— or —C≡C— groups,    -   R³ is H or A    -   m* is 1, 2, 3, 4, 5, 6, 7 or 8,    -   and    -   m is 1, 2, 3, 4, 5, 6, 7 or 8        -   wherein at least one of R^(s) and R^(b) denotes CF₃.            and pharmaceutically acceptable derivatives, solvates,            tautomers, salts and stereoisomers thereof, including            mixtures thereof in all ratios.

In another embodiment, the present invention provides compounds ofFormula (AA)

wherein R¹, R², R^(a), X, Q, S and W are as above definedand pharmaceutically acceptable derivatives, solvates, tautomers, saltsand stereoisomers thereof, including mixtures thereof in all ratios.

In another embodiment, the present invention provides compounds ofFormula (AC)

wherein R^(s) is CH₃, CH₂CH₃, F, Br, Cl, or CF₃, preferably F, CH₃ orCF₃,G₁, G₂ independently from one another denote H, Hal, or CH₃, preferablyH.And wherein R^(b), W, R¹, R², Q and S are as above defined.

In another preferred embodiment, the present invention providescompounds of Formula (AD)

Wherein G₁, G₂, R^(b), R¹, R², Q and S are as above defined.

In a very preferred embodiment, the present invention provides compoundsof Formula (AE)

wherein G₁, G₂, R^(s), R^(b), R¹, R², R³ are as defined above.Preferably G¹, G² are H.

In a more preferred embodiment, the present invention provides compoundsof Formula AF:

Wherein R^(s) is as above defined.

In a more preferred embodiment, the present invention provides compoundsof Formula AF′:

Wherein R^(s) is as above defined.

In another preferred embodiment, the present invention providescompounds of Formula AG:

Wherein R^(b) is as above defined.

In another preferred embodiment, the present invention providescompounds of Formula AG′:

Wherein R^(b) is as above defined.

Preferred embodiments of formula (I) are the compounds of formula AH,AI, AJ and AK:

wherein R^(b), R¹, R², Q and S are as above defined, and R^(s) denotesHal, A, OR³, N(R³)₂, NO₂, CN, COOR³, CF₃, OCF₃, CON(R³)₂, NR³COA,NR³CON(R³)₂, NR³SO₂A, COR³, SO₂N(R³)₂, SOA or SO₂A,

wherein R^(a), R^(b), Q, and S are as defined above,

wherein R^(a), R^(b), R¹, R², R³, X, m and m* are as defined above,

Wherein R^(a), R^(b), R¹, R², R³, W, m and m* are as defined above,

In another embodiment, the present invention provides compounds ofFormula (AL)

-   -   Wherein    -   T^(c) is an alkyl having 1 to 6 carbon atoms, preferably a CH₃        group,    -   T^(b) is CH₃, CF₃, or CH₂CH₃,    -   T^(a) is CH₃, Hal, CH₂CH₃, CF₃, and one of T^(a) and T^(b) is        CF₃.    -   d is 1, 2 or 3, preferably 1 or 2,    -   b is 1, 2 or 3 preferably 1 or 2,    -   G¹ and G² are as above defined, preferably H,    -   R³ is as above defined, preferably, H, CH₃ or CH₂CH₃,    -   G³ is one of the following groups:

The compounds of formula (I) are preferably binding on receptors forsphingosine 1-phosphate (S1P). S1P is a bioactive sphingolipidmetabolite that is secreted by hematopoietic cells and stored andreleased from activated platelets. It acts as an agonist on a family ofG protein-coupled receptors (GPCR). Five sphingosine 1-phosphatereceptors have been identified (S1P₁, S1P₂, S1P₃, S1P₄, and S1P₅, alsoknown as endothelial differentiation genes, which are Edg1, Edg5, Edg3,Edg6 and Edg8 respectively), that have widespread cellular and tissuedistribution and are well conserved in human and rodent species.

S1P is involved in a number of cellular functions such as survival,proliferation and immunological responses. The compounds of the presentinvention are preferably acting as S1P₁/Edg1 receptor agonists and thushave immunosuppressive activities by modulating leukocyte trafficking,sequestering lymphocytes in secondary lymphoid tissues, and interferingwith cell-cell interactions required for an efficient immune response.The invention is also directed to pharmaceutical compositions containingsuch compounds and methods of treatment or prevention.

FTY720 or fingolimod, a non selective S1P₁ agonist, exertsimmunosuppressive activity and shows therapeutic effects in thetreatment of relapsing-remitting multiple sclerosis. Numerouspublications have been already published using this compound: Oyster JGAnnu Rev Immunol 23:127-59, 2005, Rosen H Nat Rev Immunol 5:560-570,2005, Rosen H Trends Immunol 28:102-107, 2007, Yopp A C Clin Transplant20:788-795, 2006, Kappos L N Engl J Med 355:1124-1140, 2006, Massberg SN Engl J Med 355:1088-1089, 2006.

Immunosuppressive agents are further useful in a wide variety ofautoimmune and chronic inflammatory diseases, including systemic lupuserythematosus, chronic rheumatoid arthritis, type I diabetes mellitus,inflammatory bowel diseases, biliary cirrhosis, uveitis and otherdisorders such as Crohn's diseases, ulcerative colitis, bullouspemphigoid, sarcoidosis, psoriasis, autoimmune myositis, Wegener'sgranulomatosis, ichthyosis, Graves' ophthalmopathy, atopic dermatitisand asthma. They are also useful as part of chemotherapeutic regimensfor the treatment of cancers, lymphomas and leukemias.

Patent application WO2006/131336 describes oxadiazoles derivativescontaining a biphenyl ring. Further oxadiazole derivatives containing aphenyl group substituted with a cycloalkyl group are known from BioorgMed. Chem. Lett. 16 (2006) 3679-3683.

Oxadiazole derivatives are described in the patent applicationEP07117921.2.

It has been found that the compounds of the present invention areselective S1P₁ agonists with improved pharmacological and/or otherproperties.

The present invention uses compounds of Formula (I) and pharmaceuticallyusable derivatives, salts, tautomers, solvates and stereoisomersthereof, including mixtures thereof in all ratios, for the preparationof a medicament for the treatment and/or prophylaxis of diseases inwhich the inhibition, activation, regulation, and/or modulation of S1P₁receptor signal transduction plays a role.

Thus, the present invention preferably comprises compounds which areagonists of the S1P₁/Edg1 receptor, especially having selectivity overthe S1P₃/Edg3 receptor. An S1P₁/Edg1 receptor selective agonist hasadvantages over current therapies and extends the therapeutic window oflymphocyte sequestration agents, allowing better tolerability withhigher dosing and thus improving efficacy.

The invention further relates to the manufacture of a medicament for theimprovement of vascular function, either alone or in combination withother active compounds or therapies.

The oxadiazole compounds according to formula (I) may be prepared fromreadily available starting materials using the following general methodsand procedures. It will be appreciated that where typical or preferredexperimental conditions (i.e. reaction temperatures, time, moles ofreagents, solvents etc.) are given, other experimental conditions canalso be used unless otherwise stated. Optimum reaction conditions mayvary with the particular reactants or solvents used, but such conditionscan be determined by the person skilled in the art, using routineoptimisation procedures.

The following abbreviations refer respectively to the definitions below:

aq (aqueous), h (hour), g (gram), L (liter), mg (milligram), MHz(Megahertz), μM (micromolar) min. (minute), mm (millimeter), mmol(millimole), mM (millimolar), m.p. (melting point), eq (equivalent), mL(milliliter), μL (microliter), ACN (acetonitrile), BINAP(2,2′-bis(diphenylphosphino)-1,1′-binaphthalene, BOC(tert-butoxy-carbonyl), CBZ (carbobenzoxy), CDCl3 (deuteratedchloroform), CD3OD (deuterated methanol), CH3CN (acetonitrile), c-hex(cHex), DCC (dicyclohexyl carbodiimide), DCM (DCM), dppf(1,1′-bis(diphenylphosphino)ferrocene), DIC (diisopropyl carbodiimide),DIEA (diisopropylethyl-amine), DMF (dimethylformamide), DMSO(dimethylsulfoxide), DMSO-d6 (deuterated dimethylsulfoxide), EDC(1-(3-dimethyl-amino-propyl)-3-ethylcarbodiimide), ESI (Electro-sprayionization), EtOAc (EtOAc), Et2O (diethyl ether), EtOH (ethanol), FMOC(fluorenylmethyloxycarbonyl), HATU(dimethylamino-([1,2,3]triazolo[4,5-b]pyridin-3-yloxy)-methylene]-dimethyl-ammoniumhexafluorophosphate), HPLC (High Performance Liquid Chromatography),i-PrOH (2-propanol), K2CO3 (potassium carbonate), LC (LiquidChromatography), MD Autoprep (Mass directed Autoprep), MeOH (methanol),MgSO4 (magnesium sulfate), MS (mass spectrometry), MTBE (Methyltert-butyl ether), Mtr. (4-Methoxy-2,3,6-trimethylbenzenesulfonyl), MW(microwave), NBS (N-bromo succinimide), NaHCO3 (sodium bicarbonate),NaBH4 (sodium borohydride), NMM (N-methyl morpholine), NMR (NuclearMagnetic Resonance), POA (phenoxyacetate), Py (pyridine), PyBOP®(benzotriazole-1-yl-oxy-tris-pyrrolidino-phosphoniumhexafluorophosphate), RT (RT), Rt (retention time), SPE (solid phaseextraction), TBTU (2-(1-H-benzotriazole-1-yl)-1,1,3,3-tetramethyluromiumtetrafluoro borate), TEA (triethylamine), TFA (trifluoroacetic acid),THF (THF), TLC (Thin Layer Chromatography), UV (Ultraviolet).

Depending on the nature of R¹, R², R^(a), R^(b), W, Q, and S, differentsynthetic strategies may be selected for the synthesis of compounds offormula (I). In the process illustrated in the following schemes R¹, R²,R^(a), R^(b), W, Q, and S, are as above-defined in the description.

In general, the oxadiazole compounds according to formula (I) of thisinvention may be prepared from readily available starting materials. Ifsuch starting materials are not commercially available they may beprepared by standard synthetic techniques. The following general methodsand procedures described hereinafter in the examples may be employed toprepare compounds of formula (I). Reaction conditions depicted in thefollowing schemes, such as temperatures, solvent, or co-reagents, aregiven as examples only and are not restrictive.

Compounds of Formula (I) and related formulae, wherein R^(a), R^(b), R¹,R², Q and S are defined as above, can be converted to alternativecompounds of Formula (I) and related formulae, R^(a), R^(b), R¹, R², Qand S are defined as above, employing suitable interconversiontechniques well known by a person skilled in the art.

Generally, compounds of formula (I′), wherein R¹, R², R^(a), R^(b), Wand Q are defined as above, can be prepared by hydrolysis of the esterderivatives of formula (I″), wherein R³ is as above defined and morepreferably R³ is a methyl or tertbutyl group, using conditions wellknown to those skilled in the art, such as a metal hydroxide, e.g.lithium hydroxide, sodium hydroxide or potassium hydroxide, in asuitable solvent such as THF, methanol, ethanol or water or mixturesthereof, or using an acid, e.g. HCl or TFA, in a suitable solvent suchas dioxane, DCM, at a temperature between about 20° C. to about 50° C.,preferably at RT, for a few hours, e.g. one hour to 24 h (Scheme 1).

The method for preparing ester derivatives of Formula (I″) selectedbelow:

-   Tert-butyl    N-(3-{5-[2′-ethyl-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methyl-beta-alaninate-   Tert-butyl    N-(3-{5-[3-(methoxymethyl)-4-(2-methylpiperidin-1-yl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methyl-beta-alaninate-   Ethyl    N-(2-fluoro-4-{5-[3-(methoxymethyl)-4-(2-methylpiperidin-1-yl)phenyl]-1,2,4-oxadiazol-3-yl}benzoyl)-beta-alaninate-   Tert-butyl    [(3-{5-[3-(methoxymethyl)-4-(2-methylpiperidin-1-yl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)oxy]acetate-   Ethyl    4-(2,3-difluoro-5-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}phenoxy)butanoate-   Tert-butyl    3-[(3-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)oxy]propanoate-   Tert-butyl    3-[(3-{5-[3-(methoxymethyl)-4-(2-methylpiperidin-1-yl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)oxy]propanoate-   Tert-butyl    N-(3-{5-[2′-fluoro-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(3-{5-[2′-chloro-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    3-[(3-{5-[4-(2-methylpiperidin-1-yl)-3-(trifluoromethyl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)oxy]propanoate-   Tert-butyl    N-methyl-N-(3-{5-[4-(2-methylpiperidin-1-yl)-3-(trifluoromethyl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)glycinate-   Tert-butyl    N-methyl-N-(3-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycinate-   Methyl    2-chloro-4-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzoate

Example 13 Step 1

-   Methyl    N-(2-chloro-4-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzoyl)-beta-alaninate-   Ethyl    N-(4-{5-[2′-ethyl-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}-2-fluorobenzoyl)-beta-alaninate-   Tert-butyl    N-(3-{5-[4-(2-ethylpiperidin-1-yl)-3-(methoxymethyl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(tert-butoxycarbonyl)-N-(3-{5-[2′-ethyl-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)beta-alaninate-   Tert-butyl    N-(2-fluoro-5-{5-[3-(methoxymethyl)-4-(2-methylpiperidin-1-yl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methyl-beta-alaninate-   Tert-butyl    N-(3-fluoro-5-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(3-fluoro-5-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methyl-beta-alaninate-   Tert-butyl    N-(3-{5-[3-(methoxymethyl)-4-(2-methylpyrrolidin-1-yl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(3-{5-[3′-fluoro-2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(3-{5-[4′-fluoro-2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(3-{5-[5′-fluoro-2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    [(3-{5-[4-(2-methylpiperidin-1-yl)-3-(trifluoromethyl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)oxy]acetate-   Tert-butyl    N-(3-{5-[3′,4′-difluoro-2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Ethyl    N-[(3-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}phenyl)acetyl]-beta-alaninate-   methyl    N-[(3-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}phenyl)acetyl]-N-methylglycinate-   Tert-butyl    N-methyl-N-(3-{5-[4-[(2R)-2-methylpiperidin-1-yl]-3-(trifluoromethyl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)glycinate-   Tert-butyl    N-methyl-N-(3-{5-[4-[(2S)-2-methylpiperidin-1-yl]-3-(trifluoromethyl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)glycinate-   methyl    2,5-difluoro-4-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzoate-   Methyl    N-(2,5-difluoro-4-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzoyl)-beta-alaninate-   Tert-butyl    [(3-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)oxy]acetate-   Tert-butyl    N-(3-{5-[2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(3-{5-[4′-fluoro-2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(4-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(2-fluoro-4-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(2-bromo-4-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(4-{5-[2′-fluoro-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(2-fluoro-4-{5-[2′-fluoro-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(3-fluoro-5-{5-[4-(2-methylpiperidin-1-yl)-3-(trifluoromethyl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(3-fluoro-5-{5-[2′-fluoro-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   tert-butyl    N-isopropyl-N-(3-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycinate-   Tert-butyl    N-(tert-butoxycarbonyl)-N-(4-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycinate-   Tert-butyl    N-(tert-butoxycarbonyl)-N-(4-{5-[2′-fluoro-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycinate-   Tert-butyl    N-ethyl-N-(3-{5-[2′-fluoro-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-beta-alaninate-   Tert-butyl    N-ethyl-N-(3-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-beta-alaninate-   Tert-butyl    N-(3-fluoro-5-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(3-{5-[2′-chloro-5′-fluoro-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(2-hydroxyethyl)-N-(3-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-beta-alaninate-   Tert-butyl    N-ethyl-N-(3-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-beta-alaninate-   Tert-butyl    N-(2-fluoro-4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(2-fluoro-4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methyl-beta-alaninate-   Tert-butyl    N-(2-fluoro-4-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methyl-beta-alaninate-   Tert-butyl    N-(4-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methyl-beta-alaninate-   tert-butyl    2-((2-methoxy-4-(5-(2-(methoxymethyl)-2′-methylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)(methyl)amino)acetate-   Tert-butyl    N-methyl-N-(4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycinate-   Tert-butyl    N-(tert-butoxycarbonyl)-N-(2-fluoro-4-{5-[2′-fluoro-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycinate-   Tert-butyl    N-(tert-butoxycarbonyl)-N-(2-fluoro-4-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycinate-   Tert-butyl    N-(2-fluoro-4-{5-[4-(2-methylpiperidin-1-yl)-3-(trifluoromethyl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   tert-butyl    N-[2-(3-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}phenyl)ethyl]-N-methylglycinate-   Tert-butyl    N-(3-fluoro-5-{5-[3′-fluoro-2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(4-{5-[2′-fluoro-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(4-{5-[3′-fluoro-2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(2-fluoro-4-{5-[3′-fluoro-2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-{4-[5-(2-ethoxy-2′-methylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl]-2-fluorobenzyl}-N-methylglycinate-   Tert-butyl    N-(3-chloro-5-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(2-fluoro-4-{5-[2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(2-fluoro-3-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(4-{5-[2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(tert-oxycarbonyl)-N-(2-fluoro-4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycinate-   Tert-butyl    N-{4-[5-(2-ethoxy-2′-ethylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl]benzyl}-N-methylglycinate-   Tert-butyl    N-methyl-N-(4-{5-[2-methyl-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycinate-   Tert-butyl    N-isopropyl-N-(3-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-beta-alaninate-   Tert-butyl    N-(3-fluoro-5-{5-[2-methyl-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-isopropyl-N-(3-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-beta-alaninate-   Tert-butyl    N-(4-{5-[2′-ethyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(4-{5-[2′-ethyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}-2-fluorobenzyl)-N-methylglycinate-   Tert-butyl    N-(2-chloro-5-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(2-fluoro-3-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(2-fluoro-3-{5-[2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(4-{5-[2-ethoxy-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    4-[methyl(4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)amino]butanoate-   Tert-butyl    N-(4-{5-[2′-chloro-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(4-{5-[2′-chloro-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}-2-fluorobenzyl)-N-methylglycinate-   Tert-butyl    [(2-fluoro-4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)oxy]acetate-   Tert-butyl    N-(4-{5-[2′-chloro-3′-fluoro-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}-2-fluorobenzyl)-N-methylglycinate-   Tert-butyl    N-(4-{5-[2′-chloro-3′-fluoro-2-(methoxymethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   tert-butyl    N-(2-ethyl-4-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}phenyl)-N-methylglycinate-   Tert-butyl    N-(3,4-difluoro-5-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(4-{5-[5′-fluoro-2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   tert-butyl    2-((2-chloro-4-(5-(2-(methoxymethyl)-2′-methylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)(methyl)amino)acetate-   Tert-butyl    N-{4-[5-(2,2′-dimethylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl]benzyl}-N-methylglycinate-   Tert-butyl    N-{4-[5-(2,2′-dimethylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl]-2-fluorobenzyl}-N-methylglycinate-   Tert-butyl    N-{3-[5-(2,2′-dimethylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl]benzyl}-N-methylglycinate-   Tert-butyl    N-{3-[5-(2,2′-dimethylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl]-5-fluorobenzyl}-N-methylglycinate-   Tert-butyl    N-(4-{5-[2-chloro-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}-2-fluorobenzyl)-N-methylglycinate-   Tert-butyl    N-(4-{5-[2-chloro-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(3-{5-[2-chloro-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}-5-fluorobenzyl)-N-methylglycinate-   Tert-butyl    N-(4-{5-[2′-(difluoromethyl)-2-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    N-(2-methoxyethyl)-N-(4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycinate-   Tert-butyl    [(4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)oxy]acetate-   Tert-butyl    N-(4-{5-[2-(1-methoxyethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate-   Tert-butyl    [(4-{5-[2-(1-methoxyethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)oxy]acetate-   Tert-butyl    2-((3-chloro-4-(5-(2-(methoxymethyl)-2′-methylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)(methyl)amino)acetate-   Tert-butyl    2-((2,6-difluoro-4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)(methyl)amino)acetate-   Tert-butyl    N-(2-ethyl-4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}phenyl)-N-methylglycinate-   is more particularly described in the examples.

Alternatively, alcohol derivatives of Formula (I′″) may be convertedinto the corresponding amine derivatives of Formula (I), whereinQ=(CH₂)_(m)X(CH₂)_(m) with X=—NR³—, and S, m, R¹, R², R³, R^(a) andR^(b) are defined as above, as outlined in Scheme 1a. Compounds ofFormula (I′″) can be first be transformed into the corresponding mesylor tosyl groups (I″″), which can then reacted with an amineHN(R³)(CH₂)_(m)S, affording compounds of Formula (I) whereinQ-S═(CH₂)_(m)N(R³)(CH₂)_(m)S and S, m, R¹, R², R³, R^(a) and R^(b) aredefined as above (Scheme 1a). Alcohol (I′″) can be oxidized into thecorresponding aldehyde (I′″″), using conditions well known to thoseskilled in the art, such as but not limited to Swern oxidationconditions, or the use of MnO₂ as oxidative agent for benzylic alcohols,as illustrated on Scheme 1a. Then a reductive amination of the compoundsof Formula (I′″″) with a suitable amine HN(R³)(CH₂)_(m)S, affordscompounds of Formula (I), wherein S, m, R¹, R², R³, R^(a) and R^(b) aredefined as above.

The method for preparing alcohol derivatives of Formula (I′″) selectedbelow:

-   (3-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}phenyl)methanol-   2-(3-{5-[2-(methoxymethyl)-2′-methylbiphenyl-4-yl]-1,2,4-oxadiazol-3-yl}phenyl)ethanol-   2-(4-(5-(2-(methoxymethyl)-2′-methylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl)phenyl)ethanol-   (4-(5-(2-(methoxymethyl)-2′-methylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl)-3-methylphenyl)methanol-   (4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)phenyl)methanol-   (4-(5-(2-(methoxymethyl)-2′-methylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl)-2-methylphenyl)methanol-   (3-fluoro-4-(5-(2-(methoxymethyl)-2′-methylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl)phenyl)methanol-   (2,6-difluoro-4-(5-(2-(methoxymethyl)-2′-methylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl)phenyl)methanol-   (2-methyl-4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)phenyl)methanol    is more particularly described in the examples.

The method for preparing aldehyde derivatives of Formula (I′″″) selectedbelow:

-   4-(5-(2-(methoxymethyl)-2′-methylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl)-3-methylbenzaldehyde-   4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzaldehyde-   4-(5-(2-(methoxymethyl)-2′-methylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl)-2-methylbenzaldehyde-   is more particularly described in the examples.

The compounds of formula (I), wherein R¹, R², R^(a), R^(b), W, Q, and Sare defined as above, can be obtained in a 2-step protocol as outlinedin Scheme 2. The first step consists in the coupling of a carboxylicacid of formula (VII) with an amidoxime of formula (VI), wherein R¹, R²,R^(a), R^(b), W, Q, and S are defined as above. General protocols forsuch coupling are given below in the examples, using conditions andmethods well known to those skilled in the art to prepare anO-substituted amidoximes (V) from a carboxylic acid (VII) and an arylamidoxime (VI), with standard coupling agents, such as but not limitedto EDC, HATU, TBTU, in the presence or absence of bases such as TEA,DIEA, NMM in a suitable solvent such as DCM, ACN, THF or DMF, at atemperature rising from about 20° C. to about 50° C., preferably at RT,for a few hours, e.g. one hour to 24 h. Alternatively, a carboxylic acidderivative (e.g. acyl chloride VIIa) may be coupled with the amidoxime(VI), using conditions and methods well known to those skilled in theart, in the presence of bases such as TEA, DIEA, NMM in a suitablesolvent such as DCM, THF or DMF, at a temperature rising from about 20°C. to about 50° C., preferably at RT, for a few hours, e.g. one hour to24 h (Scheme 3). The second step consists of the cyclization anddehydration of the O-substituted amidoximes (V) to form oxadiazole (I).Conditions are given below in the examples, using methods well known tothose skilled in the art to prepare oxadiazole, such as thermolysis attemperature rising from RT to about 150° C., typically 150° C., usingpossibly a microwave oven, for a time comprised between 15 minutes and24 hours, preferably for 30 min, in a suitable solvent or mixture ofsolvents such as ACN, THF, Pyridine, DMF, in the presence or absence ofa base such as DIEA, TEA, or tetrabutyl ammonium fluoride.

Compounds of formula (VII), wherein R^(a), R^(b) and W are defined asabove, are either commercially available or may be prepared by standardsynthetic techniques, as hereinafter described in the examples, forexample by metal catalyzed coupling reaction or aromatic nucleophilicsubstitution on the corresponding halogenated benzoic acid or alkylbenzoate. Alternatively, compounds of formula (VII), wherein R^(a),R^(b) and W are defined as above, may be obtained by metal catalyzedcross-coupling reaction followed by hydrolysis of the resulting ester(XI), as shown in Scheme 4 below. More particularly, they may beobtained by Suzuki-Miyura coupling reaction between an alkyl benzoate(VIII), where R^(c) may preferably be Br, I or a sulfonate ester such astriflate, and a boronic acid (Xa) or ester (Xb), using well knownSuzuki-Miyura reaction conditions such as shown in Scheme 4 (Miyaura,N.; Suzuki, A. Chem. Rev. 1995, 95, 2457; Takahiro I. and Toshiaki M.,Tetrahedron Lett. 2005, 46, 3573-3577). In a typical procedure, alkylbenzoate (VIII) and boronic acid (Xa) or ester (Xb) are heated atvarious temperature by traditional thermic methods or using microwavetechnology in the presence of a base such as but not limited to acarbonate salt, e.g. K₂CO₃, Na₂CO₃, Cs₂CO₃, and a catalytic amount ofpalladium catalyst such as Pd(PPh₃)₄, PdCl₂(PPh₃)₂, Pd(OAc)₂, with thepossible addition of phosphine ligands such as PPh₃, S-Phos, X-Phos inan appropriate solvent or mixture of solvents such as THF, Toluene,Dioxane, MeOH, ACN, DMF, water. All the different combinations describedabove may be used. Alternatively, alkyl benzoate (IX) wherein R^(c) isas above defined and boronic acid (Xc) or ester (Xd) may be coupledunder the same palladium catalyzed procedure as described above. Theresulting ester (XI) can then be hydrolyzed using conditions well knownto those skilled in the art, such as but not limited to the use of ametal hydroxide, e.g. lithium hydroxide, sodium hydroxide or potassiumhydroxide, in a suitable solvent such as THF, methanol, ethanol or wateror mixtures thereof, at a temperature rising from about 20° C. to about60° C., preferably at RT, for a few hours, e.g. one hour to 24 h.

An alternative route for the preparation of compounds of formula (VII),wherein R^(a), R^(b) and W are defined as above, maybe via Suzuki-Miyuracoupling reaction between an alkyl benzoate boronic acid or esterderivative of formula (XII) or (XIII), where R^(d) is a boronic acid ortetramethyl-dioxaborolane, with an optionally substituted aryl,respectively (Xe) and (Xf), where R^(c) is preferably Br, I or asulfonate ester such as triflate, using well known Suzuki-Miyurareaction conditions such as shown in Scheme 5 below and described above.The resulting ester can be hydrolyzed into compounds of formula (VII)under conditions described above and in the examples below.

An alternative route for the preparation of compounds of formula (VII),wherein R^(a), R^(b) and W are defined as above, may be the addition ofan amino derivative R^(a)H of formula (Xg) to an alkyl benzoate offormula (VIIIa) or a benzoic acid of formula (VIIIb), as outlined inScheme 6, in the optional presence of a suitable base, such as TEA,DIEA, NMM in a solvent such as THF or DMF, at a temperature rising fromabout 20° C. to about 100° C., preferably at RT, for a few hours, e.g.one hour to 24 h. An amino derivative R^(a)H of formula (Xg) can be alsoused neat, as solvent. Alternatively, compounds of formula (VII) may beobtained by addition of an amino derivative R^(b)H of formula (Xh) to analkyl benzoate (IXa) or a benzoic acid (IXb), as outlined in Scheme 6,under reaction conditions described above and in the examples below. Inthe cases where ester of formula (XI) is first obtained, it can behydrolyzed into compounds of formula (VII) under conditions describedabove and in the examples below. Alternatively, an amino derivative offormula (Xg) and (Xh) can be added respectively to benzonitrile (XIVa)and (XIVb) under similar conditions as the one described above and inthe examples below. The resulting benzonitrile of formula (XIV) can behydrolyzed into the corresponding ester (XI), using conditions wellknown to those skilled in the art, such as but not limited to the use ofan acid, e.g. HCl, in a suitable solvent such as THF, methanol or wateror mixtures thereof, at a temperature rising from about 20° C. to about100° C., preferably at 78° C., for 12 h to 48 h.

Compounds of formula (VIIIa) and (IXa), wherein R^(a), R^(b) and W aredefined as above, are either commercially available or may be preparedby standard synthetic techniques, as hereinafter described in theexamples. Typically, they may be prepared by esterification of thecorresponding benzoic acid, (VIIIb) and (IXb) respectively, such as butnot limited to the formation of the corresponding acid chloride withoxalyl chloride, followed by the addition of the suitable alcohol, suchas MeOH for methyl carboxylate, at temperatures ranging from about 0° C.to about 50° C., preferably at RT for a few hours, e.g. one hour to 24hours.

Alternatively, compounds of formula (VII), wherein R^(a), R^(b) and Ware defined as above, may be obtained by metal catalyzed cross-couplingreaction followed by hydrolysis of the resulting ester of formula (XI),as shown in Scheme 7 below. More particularly, they may be obtained byBuchwald-Hartwig cross-coupling reaction between an alkyl benzoate offormula (VIII) or (IX), where R^(c) may preferably be Br, I or asulfonate ester such as triflate, and an amino derivative, respectively(Xg) or (Xh), using well known Buchwald-Hartwig reaction conditions suchas shown in Scheme 7 below (Muci, A. R.; Buchwald, S. L. Top. Curr.Chem. 2002, 219, 131-209; Jiang, L.; Buchwald, S. L. Metal-CatalyzedCross-Coupling Reactions (2^(nd) Edition) 2004, 2, 699-760). In atypical procedure, alkyl benzoate of formula (VIII) and (IX), andrespectively amino derivatives (Xg) and (Xh) are heated at varioustemperature by traditional thermic methods or using microwave technologyin presence of a base such as but not limited to a carbonate salt, e.g.K₂CO₃, Na₂CO₃, Cs₂CO₃, and a catalytic amount of palladium catalyst suchas Pd(PPh₃)₄, PdCl₂(PPh₃)₂, Pd(OAc)₂, with the possible addition ofphosphine ligands such as BINAP, X-phos, in an appropriate solvent ormixture of solvents such as THF, Toluene, Dioxane, MeOH, ACN, DMF,water. All the different combinations described above may be used. Theresulting ester of formula (XI) can be then hydrolyzed using conditionswell known to those skilled in the art, such as but not limited to theuse of a metal hydroxide, e.g. lithium hydroxide, sodium hydroxide orpotassium hydroxide, in a suitable solvent such as THF, methanol,ethanol or water or mixtures thereof, at a temperature rising from about20° C. to about 60° C., preferably at RT, for a few hours, e.g. one hourto 24 h.

Alternatively, compounds of formula (VII) wherein W is as defined aboveand wherein R^(a) or R^(b) is OA can be prepared by adding alkyl bromide(Xi) or (Xj) to the corresponding intermediates of formula (VIIIp) or(IXc) respectively, in the presence of a base, e.g K₂CO₃ in DMF at about90° C. Such transformation can also be performed on compounds of formula(I), wherein R^(a) or R^(b) is OH, as it is described hereinafter in theexamples.

Compounds of formula (VIII) are either commercially available or may beprepared by standard synthetic techniques, as hereinafter described inthe examples. Typically, when R^(b) is (CH₂)OH, (CH₂)OA, (CH₂)N(R³)₂ or(CH₂)SO₂Me, R^(c) is F, Cl, Br, I or a sulfonate ester such as triflateand R³ is as defined above, compounds of formula (VIII), respectively(VIIIf), (VIIIg), (VIIIh) and (VIIIj), may be prepared by bromination ofthe corresponding toluoyl derivative (VIIIc) followed by an S_(N)2reaction on the benzyl bromine derivative (VIIId) with a suitable group,such as but not exclusively, an acetate salt, e.g. NaOAc in HOAc, analcoholate salt, e.g. NaOA in the corresponding alcohol, THF or DMF, analcohol, e.g. HOA, that can be used as solvent, an amine, e.g. HN(R³)₂or a thiolate salt, e.g. NaSA, in a suitable solvent, such as but notexclusively THF, MeCN, DMF, at a temperature ranging from RT to 130° C.,with the possible use of the microwave (see Scheme 9). Hydrolysis of theacetate group on compounds of formula (VIIIe), using conditions wellknown to those skilled in the art, such as but not limited to sodiumhydroxide in EtOH at about 60° C., afforded compounds of formula(VIIIf). Sulfide oxidation of compounds of formula (VIIII), usingconditions well known to those skilled in the art, such as but notlimited to mCPBA, afforded compounds of formula (VIIIj). Compounds offormula (VIIIf), when Rb is (CH₂)OH, can be further transformed into thecorresponding alkyl sulfonate (VIIIk) that can be used as startingmaterial for S_(N)2 reactions similarly to (VIIIb), as it illustrated onScheme 9. Such diversification can also be performed at a later stage,on compounds of formula (I) wherein R^(b) is (CH₂)OH, as describedhereinafter in the examples.

Alternatively, compounds of formula (Miff) can be prepared by doublebromination of (VIIIc), followed by hydrolysis of (VIIIm), as it isdescribed in Scheme 10. The reduction of the resulting benzaldehydederivative of formula (VIIIn), with a suitable reducing agent, such asbut not limited to NaBH₄, yields the benzylic alcohol of (VIIIf),compound (VIIIo). Transformation of compounds of formula (VIIIn) intocompounds of formula (XIa) by metal catalyzed cross coupling reaction orS_(N)Ar reaction can be performed first. Then the reduction gives thecorresponding alcohol of formula (XIa), as outlined in Scheme 10.

When R^(b) is (CH₂)_(n)NR³SO₂A or (CH₂)_(n)NR³COA, wherein n=0 andR^(a), R³ and W are defined as above, compounds of formula (XIe) or(XIf) respectively can be synthesized from compounds of formula (XIc),as it is outlined in Scheme 11. After reduction of nitro group, theresulting aniline (XId) can be transformed into a sulphonamide (XIe)with ASO₂Cl addition or into an amide (XIf) with ACOCl addition, in thepresence of a base, such as but not limited to TEA, DIEA, NMM, pyridine,in a solvent or a mixture of solvents such as DCM, DMF, Pyridine. Suchdiversification can also be performed on a later stage, on compounds offormula (I) where R^(b) is NH₂, as it is described hereinafter in theexamples.

Compounds of formula (VIIb), where R^(b) is (CH₂)₃OA, can be synthesizedfrom compound (VIIIp) via the sequential Sonogashira and Suzuki-Miyuracross-coupling reactions, as it is outlined in Scheme 12. The resultingcompounds of formula (VIIa) can be then reduced by standard techniqueswell known to those skilled in the art, such as but not limited to Pd/Cin H₂ atmosphere, affording compounds of formula (VIIb).

Alternatively, compounds of formula (VII), wherein R^(a), R^(b) and Ware defined as above, may be prepared from compounds of formula (XV) ina two steps process, as outlined in Scheme 13. The first step is anhalogen-metal exchange with, typically but not exclusively, an alkyllithium salt, such as nBuLi or tBuLi. The second step is the addition ofCO2, at gas or solid state, as electrophile.

The method for preparing benzoic acids of Formula (VII) selected below:

-   3′-fluoro-2-(methoxymethyl)-2′-methylbiphenyl-4-carboxylic acid-   3-(methoxymethyl)-4-(2-methylpiperidin-1-yl)benzoic acid-   2-(methoxymethyl)-2′-methyl biphenyl-4-carboxylic acid-   4-(2-methylpiperidin-1-yl)-3-(trifluoromethyl)benzoic acid-   2′-methyl-2-(trifluoromethyl)biphenyl-4-carboxylic acid-   2′-ethyl-2-(methoxymethyl)-1,1′-biphenyl-4-carboxylic acid-   2-[(dimethylamino)methyl]-2′-methylbiphenyl-4-carboxylic acid-   4-(2-ethylpiperidin-1-yl)-3-(methoxymethyl)benzoic acid,-   4-[(2R)-2-methylpiperidin-1-yl]-3-(trifluoromethyl)benzoic acid-   4-[(2S)-2-methylpiperidin-1-yl]-3-(trifluoromethyl)benzoic acid-   2′-fluoro-2-(methoxymethyl)biphenyl-4-carboxylic acid-   2′-chloro-2-(methoxymethyl)biphenyl-4-carboxylic acid-   3-(methoxymethyl)-4-(2-methylpyrrolidin-1-yl)benzoic acid-   2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-carboxylic acid-   2-ethoxy-2′-methyl-1,1′-biphenyl-4-carboxylic acid-   2-methyl-2′-(trifluoromethyl)biphenyl-4-carboxylic acid-   2′-ethyl-2-(trifluoromethyl)biphenyl-4-carboxylic acid-   2,2′-dimethyl-1,1′-biphenyl-4-carboxylic acid-   2-chloro-2′-(trifluoromethyl)biphenyl-4-carboxylic acid-   2′-(difluoromethyl)-2-methylbiphenyl-4-carboxylic acid-   2′-fluoro-2-(trifluoromethyl)biphenyl-4-carboxylic acid-   is more particularly described in the examples.

Compounds of formula (VI), wherein, R¹, R², Q and S are defined asabove, can be prepared according to Scheme 14 by addition ofhydroxylamine to the corresponding substituted benzonitrile of formula(XVI) in a solvent or a mixture of solvents, such as EtOH, water, at atemperature ranging from about 20° C. to about 50° C., preferably at RT,for a few hours, e.g. one hour to 24 h.

The method for preparing amidoxime of Formula (VI) selected below:

-   ethyl    N-{-4-[amino(hydroxyimino)methyl]-2-fluorobenzoyl}-beta-alaninate-   tert-butyl ({4-[amino(hydroxyimino)methyl]benzyl}oxy)acetate-   tert-butyl    3-[{3-[amino(hydroxyimino)methyl]benzyl}methyl)amino]propanoate-   tert-yl ({3-[amino(hydroxyimino)methyl]benzyl}oxy)acetate-   ethyl 4-{5-[amino(hydroxyimino)methyl]-2,3-difluoro    phenoxy}butanoate-   N′-hydroxy-3-(methylsulfonyl)benzenecarboximidamide-   tert-butyl 3-({3-[amino(hydroxyimino)methyl]benzyl}oxy)propanoate-   tert-butyl[{3-[amino(hydroxyimino)methyl]benzyl}methyl)amino]acetate-   tert-butyl    N-{3-[amino(hydroxyimino)methyl]benzyl}-N-(tert-butoxycarbonyl)-beta-alaninate-   tert-butyl    N-{5-[amino(hydroxyimino)methyl]-2-fluorobenzyl}-N-methyl-beta-alaninate-   tert-butyl    N-{3-[amino(hydroxyimino)methyl]-5-fluorobenzyl}-N-methylglycinate-   tert-butyl    N-{3-[amino(hydroxyimino)methyl]-5-fluorobenzyl}-N-methyl-beta-alaninate-   3-[(2,3-dihydroxypropyl)amino]-N′-hydroxybenzenecarboximidamide-   N′-hydroxy-4-(2-hydroxyethoxy)benzenecarboximidamide-   4-(2-{[tert-butyl(dimethyl)silyl]oxy}ethoxy)-3-fluoro-N′-hydroxybenzenecarboximidamide-   3-{[(2,3-dihydroxypropyl)(methyl)amino]methyl}-N′-hydroxybenzenecarboximidamide-   3-{[bis(2-hydroxyethyl)amino]methyl}-N′-hydroxybenzenecarboximidamide-   tert-butyl N-{4-[amino(hydroxyimino)methyl]benzyl}-N-methylglycinate-   tert-butyl    N-{-4-[amino(hydroxyimino)methyl]-2-fluorobenzyl}-N-methylglycinate-   tert-butyl    N-{-4-[amino(hydroxyimino)methyl]-2-bromobenzyl}-N-methylglycinate-   methyl 4-[amino(hydroxyimino)methyl]-2-chlorobenzoate-   methyl 4-[amino(hydroxyimino)methyl]-2-fluorobenzoate-   tert-butyl    N-{4-[amino(hydroxyimino)methyl]benzyl}-N-(tert-butoxycarbonyl)glycinate-   3-{ethyl-[3-(n-hydroxycarbamimidoyl)-benzyl]-amino}-propionic acid    tert-butyl ester-   tert-butyl    N-{3-[amino(hydroxyimino)methyl]benzyl}-N-(2-hydroxyethyl)-beta-alaninate-   tert-butyl    N-{4-[amino(hydroxyimino)methyl]-2-fluorobenzyl}-N-methyl-beta-alaninate-   tert-butyl    N-{4-[amino(hydroxyimino)methyl]benzyl}-N-methyl-beta-alaninate-   methyl 4-[amino(hydroxyimino)methyl]-2,5-difluorobenzoate-   tert-butyl    N-{-4-[amino(hydroxyimino)methyl]-2-methoxybenzyl}-N-methylglycinate-   tert-butyl    N-{-4-[amino(hydroxyimino)methyl]-2-fluorobenzyl}-N-(tert-butoxycarbonyl)glycinate-   tert-butyl    N-{2-fluoro-3-[(hydroxyamino)(imino)methyl]benzyl}-N-methylglycinate-   3-{[3-(N-Hydroxycarbamimidoyl)-benzyl]-isopropyl-amino}-propionic    acid tert-butyl ester-   tert-butyl    N-{5-[amino(hydroxyimino)methyl]-2-chlorobenzyl}-N-methylglycinate-   tert-butyl4-[{4-[amino(hydroxyimino)methyl]benzyl}(methyl)amino]butanoate-   tert-butyl    2-((3-chloro-4-(N′-hydroxycarbamimidoyl)benzyl)(methyl)amino)acetate-   N′-hydroxy-4-(hydroxymethyl)-3-methylbenzimidamide-   tert-butyl    2-((2-ethyl-4-(N′-hydroxycarbamimidoyl)benzyl)(methyl)amino)acetate-   N′-hydroxy-3-(hydroxymethyl)benzimidamide-   tert-butyl    N-{4-[amino(hydroxyimino)methyl]benzyl}-N-(2-methoxyethyl)glycinate-   N′-hydroxy-4-(hydroxymethyl)benzenecarboximidamide-   tert-butyl    2-((3-chloro-4-(N′-hydroxycarbamimidoyl)benzyl)(methyl)amino)acetate:-   tert-butyl    2-((2,6-difluoro-4-(N′-hydroxycarbamimidoyl)benzyl)(methyl)amino)acetate-   {3-[amino(hydroxyimino)methyl]phenyl}acetic acid-   is more particularly described in the examples.

Compounds of formula (XVI), wherein R¹, R², Q and S are defined asabove, are either commercially available or may be prepared by standardsynthetic techniques well known to those skilled in the art. Typically,when S is COOR³ as defined above, compounds of formula (XVIa) may beprepared by coupling the corresponding carboxylic acid of formula (XVII)to an alcohol, using conditions such as but not limited to the formationof the corresponding acid chloride of compounds of formula (XVII) withoxalyl chloride, followed by the addition of the suitable alcohol, suchas MeOH at temperatures ranging from about 0° C. to about 50° C.,preferably at RT for few hours, e.g. one hour to 24 hours, as it isoutlined on Scheme 15. It may be also prepared with standard couplingagents, such as but not limited to EDC, HATU, TBTU, in the presence orabsence of bases such as TEA, DIEA, NMM in the suitable alcohol, such asMeOH, at a temperature between about 20° C. to about 50° C., preferablyat RT, for a few hours, e.g. one hour to 24 h.

Alternatively compounds of formula (XVIb) wherein Q=(CH₂)_(m)X(CH₂)_(m),X═—O—, and R¹, R², m and S are defined as above, may be prepared fromalcohol (XVIII), by addition of an electrophile, LG-(CH₂)_(m)S, whereLG- is a leaving group, such as but not exclusively Br, I, OMs, in thepresence of a base such as LiHMDS, NaH, NaOH, in a solvent or a mixtureof solvent such as THF or Toluene-water in the presence of a phasetransfer agent, such as but not limited to (Bu₄N)HSO₄ at temperaturesranging from RT to about 100° C., as it is outlined in Scheme 16.Alternatively, alcohol (XVIII) can be transformed into the correspondingmesyl or tosyl groups, which can then react with an alcohol (for X═—O—)or an amine (for X═—NR³—), affording compounds of formula (XVIb) and(XVIc) respectively, wherein Q=(CH₂)_(m)X(CH₂)_(m) and R¹, R², m and Sare defined as above (Scheme 16). Alcohol (XVIII) can be oxidized intothe corresponding aldehyde (XVIIIb), according to Scheme 16.

Then a reductive amination of the compounds of formula (XVIIIb) with asuitable amine, affords compounds of formula (XVIc), whereinQ=(CH₂)_(m)X(CH₂)_(m) with X═—NR³—, and R¹, R², m and S are defined asabove, according to Scheme 16. The different transformations describedin Scheme 16 may be performed on compounds of formula (I) with thesuitable substitution pattern, as it is described in the examples.

Alternatively, addition of an alcohol or an amine to benzyl bromide offormula (XVIIIc), as outlined in Scheme 17, in the presence of a base,such as but not limited to DIEA, TEA, K₂CO₃, Cs₂CO₃, in a suitablesolvent such as MeCN, THF, DMF, yields compounds of formula (XVIb) and(XVIc), wherein Q=(CH₂)X(CH₂)_(m), X═—O— or —NR³— respectively, and R¹,R², m and S are defined as above.

Phenol derivative of formula (XIX) may be transformed into compounds offormula (XVId) by Mitsunobu or alkylation reaction, using conditionsknown to the person skilled in the art and as described below in theexamples. Typically, phenol alkylation with LG-(CH₂)_(m)S, where LG- isa leaving group, such as but not limited to Br, I, OMs, is performed ina solvent such as THF or DMF, in the presence of a base such as DIEA,TEA, K₂CO₃ or Cs₂CO₃, at temperature ranging from RT to about 100° C.

Alternatively, aniline derivative of formula (XX) may be transformedinto compounds of formula (XVIe) by alkylation reaction, usingconditions known to the person skilled in the art and as described belowin Scheme 19 and in the examples. Typically, aniline alkylation withLG-(CH₂)_(m)S, where LG- is a leaving group, such as but not exclusivelyBr, I, OMs, is performed in a solvent such as THF or DMF, in thepresence of a base such as DIEA, TEA, K₂CO₃ or Cs₂CO₃, at temperatureranging from RT to about 100° C.

Alternatively, compounds of formula (XVI), wherein Q, S, R¹, and R² aredefined as above, may be obtained from the corresponding aryl fluoride(XIII) by aromatic nucleophilic substitution with a cyanide salt,typically but not exclusively sodium cyanide in the presence oftetrabutylammonium bromide, in a solvent such as DMF and at atemperature ranging from about 20° C. to about 100° C., preferably atabout 60° C., for few hours, e.g. 12 h, as it is described in Scheme 20,according to Jenkins, T. J. et al. J. Med. Chem. 2007, 50, 566.

Metal catalyzed cyanation of aryl bromide of formula (XIV) can be usedas alternative strategy, as shown on Scheme 20. Addition of Zn(CN)₂ inthe presence of a palladium catalyst, such as but not limited toPd₂(dba)₃ or Pd(PPh₃)₄, with the optional addition of a ligand such asdppf (according to Maligres, P. E. et al Tetrahedron Lett. 1999, 40,8193-8195), and zinc derivatives such as but not limited to Zn dust andZn(OAc)₂ (according to Chidambaram, R. et al Tetrahedron Lett. 2004, 45,1441-1444) in a solvent such as DMF and at temperature raising from RTto 150° C., typically 100° C., yields the formation of compounds offormula (XI). The cyanation of aryl bromide of formula (XIV) can be alsoperformed in the absence of palladium, with the use of CuCN in DMF(according to Couture. C.; Paine, A. J. Can. J. Chem. 1985, 63,111-120).

Cyanide group may be introduced by Sandmeyer reaction, starting from ananiline of formula (XXI), as outlined in Scheme 20. Its transformationinto the corresponding diazonium salt can be achieved with sodiumnitrite in the presence of a mineral acid, such as HCl in water. It canthen further react with copper cyanide, prepared from a mixture of CuCNand KCN, in water at a temperature ranging from about 20° C. to about100° C., affording compounds of formula (XI) (according to Barraclough,P. et al. Arch. Pharm. 1990, 323, 507-512). The starting anlinederivatives of formula (XXI) are either commercially available or can beobtained by reduction of the corresponding nitro group by Pd/C catalyzedhydrogenation, as described hereafter in the examples.

If the above set out general synthetic methods are not applicable toobtain the compounds of formula (I), suitable methods of preparationknown by a person skilled in the art should be used.

The pharmaceutically acceptable cationic salts of compounds of thepresent invention are readily prepared by reacting the acid forms withan appropriate base, usually one equivalent, in a co-solvent. Typicalbases are sodium hydroxide, sodium methoxide, sodium ethoxide, sodiumhydride, potassium hydroxide, potassium methoxide, magnesium hydroxide,calcium hydroxide, benzathine, choline, diethanolamine, ethylenediamine,meglumine, benethamine, diethylamine, piperazine and tromethamine. Thesalt is isolated by concentration to dryness or by addition of anon-solvent. In some cases, salts can be prepared by mixing a solutionof the acid with a solution of the cation (sodium ethylhexanoate,magnesium oleate), employing a solvent in which the desired cationicsalt precipitates, or can be otherwise isolated by concentration andaddition of a non-solvent.

According to a further general process, compounds of formula (I), andany subformulae can be converted to alternative compounds of formula (I)and any subformulae, employing suitable inter-conversion techniques wellknown by a person skilled in the art.

In general, the synthesis pathways for any individual compounds offormula (I) will depend on the specific substituents of each moleculeand upon the ready availability of Intermediates necessary; again suchfactors being appreciated by those of ordinary skill in the art. For allthe protection and de-protection methods, see Philip J. Kocienski, in“Protecting Groups”, Georg Thieme Verlag Stuttgart, New York, 1994 and,Theodora W. Greene and Peter G. M. Wuts in “Protective Groups in OrganicSynthesis”, Wiley Interscience, 3^(rd) Edition 1999.

Compounds of this invention can be isolated in association with solventmolecules by crystallization from evaporation of an appropriate solvent.The pharmaceutically acceptable acid addition salts of the compounds offormula (I), which contain a basic center, may be prepared in aconventional manner. For example, a solution of the free base may betreated with a suitable acid, either neat or in a suitable solution, andthe resulting salt isolated either by filtration or by evaporation undervacuum of the reaction solvent. Pharmaceutically acceptable baseaddition salts may be obtained in an analogous manner by treating asolution of compounds of formula (I), which contain an acid center, witha suitable base. Both types of salts may be formed or interconvertedusing ion-exchange resin techniques.

Depending on the conditions used, the reaction times are generallybetween a few minutes and 14 days, and the reaction temperature isbetween about −30° C. and 140° C., normally between −10° C. and 90° C.,in particular between about 0° C. and about 70° C.

Compounds of the formula (I) can furthermore be obtained by liberatingcompounds of the formula (I) from one of their functional derivatives bytreatment with a solvolysing or hydrogenolysing agent.

Preferred starting materials for the solvolysis or hydrogenolysis arethose which conform to the formula (I), but contain correspondingprotected amino and/or hydroxyl groups instead of one or more free aminoand/or hydroxyl groups, preferably those which carry an amino-protectinggroup instead of an H atom bound to an N atom, in particular those whichcarry an R′—N group, in which R′ denotes an amino-protecting group,instead of an HN group, and/or those which carry a hydroxyl-protectinggroup instead of the H atom of a hydroxyl group, for example those whichconform to the formula (I), but carry a —COOR″ group, in which R″denotes a hydroxyl protecting group, instead of a —COOH group.

It is also possible for a plurality of—identical or different—protectedamino and/or hydroxyl groups to be present in the molecule of thestarting material. If the protecting groups present are different fromone another, they can in many cases be cleaved off selectively.

The term “amino-protecting group” is known in general terms and relatesto groups which are suitable for protecting (blocking) an amino groupagainst chemical reactions, but which are easy to remove after thedesired chemical reaction has been carried out elsewhere in themolecule. Typical of such groups are, in particular, unsubstituted orsubstituted acyl, aryl, aralkoxymethyl or aralkyl groups. Since theamino-protecting groups are removed after the desired reaction (orreaction sequence), their type and size are furthermore not crucial;however, preference is given to those having 1-20, in particular 1-8,carbon atoms. The term “acyl group” is to be understood in the broadestsense in connection with the present process. It includes acyl groupsderived from aliphatic, araliphatic, aromatic or heterocyclic carboxylicacids or sulfonic acids, and, in particular, alkoxy-carbonyl,aryloxycarbonyl and especially aralkoxycarbonyl groups. Examples of suchacyl groups are alkanoyl, such as acetyl, propionyl and butyryl;aralkanoyl, such as phenylacetyl; aroyl, such as benzoyl and tolyl;aryloxyalkanoyl, such as POA; alkoxycarbonyl, such as methoxy-carbonyl,ethoxycarbonyl, 2,2,2-trichloroethoxycarbonyl, BOC(tert-butoxy-carbonyl) and 2-iodoethoxycarbonyl; aralkoxycarbonyl, suchas CBZ (“carbo-benz-oxy”), 4-methoxybenzyloxycarbonyl and FMOC; andaryl-sulfonyl, such as Mtr. Preferred amino-protecting groups are BOCand Mtr, furthermore CBZ, Fmoc, benzyl and acetyl.

The term “hydroxyl-protecting group” is likewise known in general termsand relates to groups which are suitable for protecting a hydroxyl groupagainst chemical reactions, but are easy to remove after the desiredchemical reaction has been carried out elsewhere in the molecule.Typical of such groups are the above-mentioned unsubstituted orsubstituted aryl, aralkyl or acyl groups, furthermore also alkyl groups.The nature and size of the hydroxyl-protecting groups are not crucialsince they are removed again after the desired chemical reaction orreaction sequence; preference is given to groups having 1-20, inparticular 1-10, carbon atoms. Examples of hydroxyl-protecting groupsare, inter alia, benzyl, 4-methoxybenzyl, p-nitro-benzoyl,p-toluenesulfonyl, tert-butyl and acetyl, where benzyl and tert-butylare particularly preferred.

The compounds of the formula (I) are liberated from their functionalderivatives—depending on the protecting group used—for example usingstrong acids, advantageously using TFA or perchloric acid, but alsousing other strong inorganic acids, such as hydrochloric acid orsulfuric acid, strong organic carboxylic acids, such as trichloroaceticacid, or sulfonic acids, such as benzene- or p-toluenesulfonic acid. Thepresence of an additional inert solvent is possible, but is not alwaysnecessary. Suitable inert solvents are preferably organic, for examplecarboxylic acids, such as acetic acid, ethers, such as THF or dioxane,amides, such as DMF, halogenated hydrocarbons, such as DCM, furthermorealso alcohols, such as methanol, ethanol or isopropanol, and water.Mixtures of the above-mentioned solvents are furthermore suitable. TFAis preferably used in excess without addition of a further solvent, andperchloric acid is preferably used in the form of a mixture of aceticacid and 70% perchloric acid in the ratio 9:1. The reaction temperaturesfor the cleavage are advantageously between about 0 and about 50° C.,preferably between 15 and 30° C. (RT).

The BOC, OBut and Mtr groups can, for example, preferably be cleaved offusing TFA in DCM or using approximately 3 to 5N HCl in dioxane at 15-30°C., and the FMOC group can be cleaved off using an approximately 5 to50% solution of dimethylamine, diethylamine or piperidine in DMF at15-30° C.

Protecting groups which can be removed hydrogenolytically (for exampleCBZ, benzyl or the liberation of the amidino group from the oxadiazolederivative thereof) can be cleaved off, for example, by treatment withhydrogen in the presence of a catalyst (for example a noble-metalcatalyst, such as palladium, advantageously on a support, such ascarbon). Suitable solvents here are those indicated above, inparticular, for example, alcohols, such as methanol or ethanol, oramides, such as DMF. The hydrogenolysis is generally carried out attemperatures between about 0 and 100° C. and pressures between about 1and 200 bar, preferably at 20-30° C. and 1-10 bar. Hydrogenolysis of theCBZ group succeeds well, for example, on 5 to 10% Pd/C in methanol orusing ammonium formate (instead of hydrogen) on Pd/C in methanol/DMF at20-30° C.

Examples of suitable inert solvents are hydrocarbons, such as hexane,petroleum ether, benzene, toluene or xylene; chlorinated hydrocarbons,such as trichloroethylene, 1,2-dichloroethane, tetrachloromethane,tri-fluoro-methylbenzene, chloroform or DCM; alcohols, such as methanol,ethanol, isopropanol, n-propanol, n-butanol or tert-butanol; ethers,such as diethyl ether, diisopropyl ether, tetrahydrofurane (THF) ordioxane; glycol ethers, such as ethylene glycol monomethyl or monoethylether or ethylene glycol dimethyl ether (diglyme); ketones, such asacetone or butanone; amides, such as acetamide, dimethylacetamide,N-methylpyrrolidone (NMP) or dimethyl-formamide (DMF); nitriles, such asacetonitrile; sulfoxides, such as dimethyl sulfoxide (DMSO); carbondisulfide; carboxylic acids, such as formic acid or acetic acid; nitrocompounds, such as nitromethane or nitrobenzene; esters, such as EtOAc,or mixtures of the said solvents.

Esters can be saponified, for example, using LiOH, NaOH or KOH in water,water/THF, water/THF/ethanol or water/dioxane, at temperatures between 0and 100° C. Furthermore, ester can be hydrolysed, for example, usingacetic acid, TFA or HCL.

Free amino groups can furthermore be acylated in a conventional mannerusing an acid chloride or anhydride or alkylated using an unsubstitutedor substituted alkyl halide or reacted with CH₃—C(═NH)—OEt,advantageously in an inert solvent, such as DCM or THF and/or in thepresence of a base, such as triethylamine or pyridine, at temperaturesbetween −60° C. and +30° C.

Therefore, the invention also relates to the preparation of thecompounds of formula (I), and salts thereof, characterized in that

a compounds of formula A:

wherein W, R^(a) and R^(b) have the meanings given above, and T is OH,or a leaving group, such as Cl, Br, I, imidazolyl, pentafluorophenoxy orthe product of the reaction of isobutyl chloroformate with formula A,wherein T is OH, is reacted witha compounds of formula B:

wherein R¹ and R² Q and S have the meanings given above preferably inthe presence of a solvent and of a suitable base, such as an amine likeTEA, DIEA or NMM, or in case T is OH, in the presence of a suitablecondensation reagent, such as EDC, HATU, and the resulting product iscyclized, preferably in the presence of an amine, such as DIEA, TEA ortetrabatylaminonium fluoride,and optionally a base or acid of the formula I is converted into one ofits salts.

Throughout the specification, the term leaving group preferably denotesCl, Br, I or a reactively modified OH group, such as, for example, anactivated ester, an imidazolide or alkylsulfonyloxy having 1-6 carbonatoms (preferably methylsulfonyloxy or trifluoromethylsulfonyloxy) orarylsulfonyloxy having 6-10 carbon atoms (preferably phenyl- orp-tolylsulfonyloxy). Radicals of this type for activation of thecarboxyl group in typical acylation reactions are described in theliterature (for example in the standard works, such as Houben-Weyl,Methoden der organischen Chemie [Methods of Organic Chemistry],Georg-Thieme-Verlag, Stuttgart). Activated esters are advantageouslyformed in situ, for example through addition of HOBt orN-hydroxysuccinimide.

The formula (I) also encompasses the optically active forms(stereoisomers), the enantiomers, the racemates, the diastereomers andthe hydrates and solvates of these compounds. The term “solvates of thecompounds” is taken to mean adductions of inert solvent molecules ontothe compounds which form owing to their mutual attractive force.Solvates are, for example, mono- or dihydrates or alcoholates.

The term “pharmaceutically usable derivatives” is taken to mean, forexample, the salts of the compounds of the formula I and so-calledprodrug compounds.

The term “prodrug derivatives” is taken to mean compounds of the formulaI which have been modified with, for example, alkyl or acyl groups,sugars or oligopeptides and which are rapidly cleaved in the organism toform the active compounds.

These also include biodegradable polymer derivatives of the compoundsaccording to the invention, as described, for example, in Int. J. Pharm.115, 61-67 (1995).

The formula (I) also encompasses mixtures of the compounds of theformula I, for example mixtures of two diastereomers, for example in theratio 1:1, 1:2, 1:3, 1:4, 1:5, 1:10, 1:100 or 1:1000.

These are particularly preferably mixtures of stereoisomeric compounds.

In a preferred embodiment, the invention relates to compounds of Formula(I) wherein R¹, R², W, Q, S and R^(b) are as defined above, and whereinR^(a) is Ar monosubstituted at the carbon adjacent to the carbon linkedto the rest of the molecule.

In another preferred embodiment, the invention relates to compounds ofFormula (I) wherein Q is on meta position with regards to the oxadiazolemoiety.

In another preferred embodiment, the present invention providescompounds of Formula (I) wherein R^(b) is CF₃ and R^(s) is CH₃ orCH₂OCH₃.

In another preferred embodiment, the present invention providescompounds of Formula (I) wherein R^(s) is CF₃ and R^(b) is CH₃ orCH₂OCH₃.

In another preferred embodiment, the present invention providescompounds of Formula (I) wherein R³ is CH₃.

In another preferred embodiment, the present invention providescompounds of Formula (I) wherein R³ is CH₃, R^(s) is CF₃ and R^(b) isCH₃ or CH₂OCH₃, or wherein R³ is CH₃, R^(b) is CF₃ and R^(s) is CH₃ orCH₂OCH₃.

In another preferred embodiment, compounds of Formula (I) exhibit aselectivity on S₁P₁ receptor over S₁P₃ receptor of more than 20 fold,preferably more than 50 fold, more preferably more than 100 fold, evenmore preferably more than 1000 fold.

The preferred compounds of the present invention have a high oralbioavailability and/or a low clearance. More particularly, compounds ofthe present invention are characterized by a ratio Cl/F, wherein Cl isthe clearance and F the bioavailability, of 0.5 or lower, morepreferably lower than 0.4, and most preferably of 0.3 or lower.

In another preferred embodiment, compounds of the present invention havea Plasma Area Under Curve (AUC∞) of 80000 h*ng/ml or higher, morepreferably higher than 100000 h*ng/ml.

In another preferred embodiment, compounds of the present inventionexhibit a selectivity on S₁P₁ receptor over S₁P₃ receptor of more than20 fold, preferably more than 50 fold, more preferably more than 100fold, even more preferably more than 1000 fold and show a ratio Cl/F,wherein Cl is the clearance and F the bioavailability, of 0.5 or lower,more preferably lower than 0.4, and most preferably of 0.3 or lower.

In another preferred embodiment, compounds of the present inventionexhibit a selectivity on S₁P₁ receptor over S₁P₃ receptor of more than20 fold, preferably more than 50 fold, more preferably more than 100fold, even more preferably more than 1000 fold and have a Plasma AreaUnder Curve (AUC) of 80000 h*ng/ml or higher, more preferably higherthan 100000 h*ng/ml.

Preference is given to the compounds of the present invention selectedfrom the following examples 1 to 155:

Example Nb structure 14

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and pharmaceutically usable derivatives, solvates, salts andstereoisomers thereof, including mixtures thereof in all ratios.

For all radicals and indices such as m which occur more than once withinthe same chemical structure, their meanings are independent of oneanother.

Above and below, the radicals or parameters R^(a), R^(b), R¹, R², R³, W,Q, S, T, X, X¹, X², A, Ar, Het, m and n have the meaning indicated underthe formula (I) and subformulae, unless expressly stated otherwise.

A denotes alkyl, is unbranched (linear) or branched, and has 1, 2, 3, 4,5, 6, 7, 8, 9, 10, 11 or 12 carbon atoms. A preferably denotes methyl,furthermore ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl ortert-butyl, furthermore also pentyl, 1-, 2- or 3-methylbutyl, 1,1-, 1,2-or 2,2-dimethylpropyl, 1-ethylpropyl, hexyl, 1-, 2-, 3- or4-methylpentyl, 1,1-, 1,2-, 1,3-, 2,2-, 2,3- or 3,3-dimethylbutyl, 1- or2-ethylbutyl, 1-ethyl-1-methylpropyl, 1-ethyl-2-methylpropyl, 1,1,2- or1,2,2-trimethylpropyl, furthermore preferably, for example,trifluoromethyl.

A very particularly preferably denotes alkyl having 1, 2, 3, 4, 5 or 6carbon atoms, preferably methyl, ethyl, propyl, isopropyl, butyl,isobutyl, sec-butyl, tert-butyl, pentyl, hexyl, trifluoromethyl,pentafluoroethyl or 1,1,1-trifluoroethyl. A furthermore denotes(CH₂)_(n)O(CH₂)_(n)OR³, (CH₂)_(n)NR³(CH₂)₂N(R³)₂, especially(CH₂)₂O(CH₂)₂OR³ or (CH₂)₂NH(CH₂)₂N(R³)₂.

Cycloalkyl is a cyclic alkyl containing 3 to 12 carbon atoms. Cycloalkylpreferably denotes cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl orcycloheptyl.

Cycloalkylalkylene is a cycloalkyl group bond to the rest of themolecule via a carbon chain and having 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,11, 12, 13, 14, 15, 16, 17, or 18 carbon atoms. Cycloalkylalkylenepreferably denotes cyclopropylmethylene, cyclobutylmethylene,cyclopentylmethylene, cyclohexylmethylene or cycloheptylmethylene.

Alkylene is a bivalent carbon chain having 1, 2, 3, 4, 5, 6, 7, 8, 9,10, 11, or 12 carbon atoms. Alkylene is preferably methylene, ethylene,propylene, butylene, pentylene or hexylene, furthermore branchedalkylene.

R^(a) is preferably Ar.

Ar is preferably substituted with CF₃, methyl, trifluoromethyl methoxyor NO₂.

If Het denotes a N-Atom bearing saturated heterocycle, Het is preferablylinked to the rest of the molecule via an N-Atom. The alpha position isnext to this N-Atom.

R^(a) very preferably denotes one of the following groups:

R^(b) is preferably H, A, OR³, CF₃, NO₂, NH₃, Hal, CH₂OR³, (CH₂)_(m)OA,especially CH₂OCH₃, CH₂NHSO₂A, NHSO₂A, such as NHSO₂CH₃, CH₂NHCOCH₃,CH₂N(CH₃)₂, CH₂NH₂, NHCONH₂ or CF₃. Very preferably, R^(b) is one of thefollowing groups:

—CH₃, —OH, NO₂, —CH₂OH, —CH₂OCH₃, —CH₂OC₂H₅, —CH₂OCH(CH₃)₂, —CH₂NHCH₃,—CH(CH₃)OCH₃, —CH₂N(CH₃)₂, —CH₂SO₂CH₃, —(CH₂)₃OCH₃, —OCH₃, —O(CH₂)₂OCH₃,—OCH₂CH(CH₃)₂, —CF₃, CN, —NHCOCH₃, —NHCOC₂H₅, —NHSO₂CH₃, —NHSO₂C₂H₅,—NHSO₂C₃H₇, —NHSO₂N(CH₃)₂, Cl,

R³ preferably denotes H, (C₁-C₆ alkyl), (C₁-C₆ fluoroalkyl), morepreferably, R³ is H. 2 geminal groups R³ linked to a N atom particularlydenote

wherein n is 0, 1, 2 or 3.

Hal is preferably F, Cl or Br and especially F or Cl.

Preferably, at least one of R¹ and R² denotes F or Cl.

R¹ preferably denotes F or O-alkyl, especially F or OCH₃,

R² is preferably H.

W preferably denotes CH.

Q is preferably in para-position, or in meta-position with respect tothe oxadiazole moiety.

S is preferably COOR₃ and especially COOH.

The group Q-S is preferably selected from the following groups:

Alternatively, the group Q-S denotes —CONH(CH₂)_(n)COOH,—CH₂CONH(CH₂)_(n)COOH, CH₂CON(CH₃)(CH₂)_(n)COOH,—CH₂CONH(CH₂)_(n)NHCOCH₃, —CH₂NH(CH₂)_(n)COOH, —CH₂N(CH₃)(CH₂)_(n)COOH,—(CH₂)₃N(CH₃)(CH₂)_(n)COOtBu, —CH₂N(iPr)(CH₂)_(n)COOH,—CH₂N(iBu)(CH₂)_(n)COOH, —CH₂N(C₂H₄OH)(CH₂)_(n)COOH,—CH₂N(C₂H₅)(CH₂)_(n)COOH, —CH₂N(C₃H₇)(CH₂)_(n)COOH,—CH₂N(CH₃)(CH₂)CH(OH)CH₂OH, —CH₂N(C₂H₄OH)₂, —CH₂CONH(CH₂)_(n)N(CH₃)₂,—SO₂Me, —O(CH₂)_(n)COOH, —O(CH₂)_(n)OH, —CH₂—O—(CH₂)_(n)COOH,NHCH₂CH(OH)CH₂OH, CONHCH₂CH(OH)CH₂OH, —CONHCH₂CH(CH₃)COOEt,—CONHCH₂CH(CH₃)COOH,

n is preferably 0, 1, 2, 3, 4 or 5 and more preferably 0, 1, 2, 3 or 4.

m is preferably 1, 2 or 3, most preferably 1 or 2,

m* is preferably 1, 2 or 3, most preferably 1 or 2.

An aromatic carbocyclic ring preferably denotes phenyl, naphthyl orbiphenyl.

Ar denotes, for example, phenyl, o-, m- or p-tolyl, o-, m- orp-ethylphenyl, o-, m- or p-propylphenyl, o-, m- or p-isopropylphenyl,o-, m- or p-tert-butylphenyl, o-, m- or p-hydroxyphenyl, o-, m- orp-nitrophenyl, o-, m- or p-aminophenyl, o-, m- orp-(N-methylamino)phenyl, o-, m- or p-(N-methylaminocarbonyl)phenyl, o-,m- or p-acetamidophenyl, o-, m- or p-methoxyphenyl, o-, m- orp-ethoxyphenyl, o-, m- or p-ethoxycarbonylphenyl, o-, m- orp-(N,N-dimethylamino)phenyl, o-, m- orp-(N,N-dimethylaminocarbonyl)phenyl, o-, m- or p-(N-ethylamino)phenyl,o-, m- or p-(N,N-diethylamino)phenyl, o-, m- or p-fluorophenyl, o-, m-or p-bromophenyl, o-, m- or p-chlorophenyl, o-, m- orp-(methylsulfonamido)phenyl, o-, m- or p-(methylsulfonyl)phenyl, o, m orp amino-sulfanyl-phenyl, o-, m- or p-phenoxyphenyl, further preferably2,3-, 2,4-, 2,5-, 2,6-, 3,4- or 3,5-dimethylphenyl, 2,3-, 2,4-, 2,5-,2,6-, 3,4- or 3,5-di-fluorophenyl, 2,3-, 2,4-, 2,5-, 2,6-, 3,4- or3,5-dichlorophenyl, 2,3-, 2,4-, 2,5-, 2,6-, 3,4- or 3,5-di-bromophenyl,2,4- or 2,5-dinitrophenyl, 2,5- or 3,4-dimethoxyphenyl,3-nitro-4-chlorophenyl, 3-amino-4-chloro-, 2-amino-3-chloro-,2-amino-4-chloro-, 2-amino-5-chloro- or 2-amino-6-chloro-phenyl,2-nitro-4-N,N-dimethylamino- or 3-nitro-4-N,N-dimethylaminophenyl,2,3-diaminophenyl, 2,3,4-, 2,3,5-, 2,3,6-, 2,4,6- or3,4,5-trichlorophenyl, 2,4,6-trimethoxyphenyl,2-hydroxy-3,5-dichlorophenyl, p-iodophenyl, 3,6-dichloro-4-aminophenyl,4-fluoro-3-chlorophenyl, 2-fluoro-4-bromophenyl,2,5-difluoro-4-bromophenyl, 3-bromo-6-methoxyphenyl,3-chloro-6-methoxy-phenyl, 3-chloro-4-acetamidophenyl,3-fluoro-4-methoxyphenyl, 3-amino-6-methylphenyl,3-chloro-4-acetamidophenyl or 2,5-dimethyl-4-chlorophenyl.

Ar preferably denotes, for example, phenyl which is unsubstituted ormonosubstituted, disubstituted or trisubstituted by A, Hal, OR³, CF₃,OCF₃, NO₂ and/or CN. If Ar is phenyl, it is preferably substituted inortho-position to the C-atom linking Ar to the rest of the molecule. Theortho-position is also indicated by the figure “2” in chemicalnomenclature. Ar is preferably substituted by, —CH₃, —(CH₂)_(n)OR³,—(CH₂)_(n)NR³SO₂A.

Ar particularly preferably denotes, for example, phenyl which isunsubstituted or monosubstituted or disubstituted preferablymonosubstituted, by OCH₃, OH, CH₃, CF₃, such as, for example,2′-methoxy-phenyl-, 2′-trifluoromethyl-phenyl- preferably, aryl bearingat least a 2′ substituent, 2′-chloro-phenyl, 2′,6′-dimethyl-phenyl- or2′-alkyl-phenyl-, preferably 2′-methyl-phenyl-.

Ar very particularly preferably denotes one of the following groups:

preferably

wherein X¹, and X² denote independently of one another F, Cl, —OCH₃,—CH₃, —C₂H₅, —CF₃, —OCF₃, —O-isoPropyl, —O-isobutyl, —OCH₂CN,—OCH₂cyclopropyl, —CH₂OH, —CH₂O-isoPropyl, —CH₂O-isobutyl,—CH₂OCH₂cyclopropyl, —CH₂NMe₂, —CH₂OC₂H₅, —NHCOMe, —NHCOEt, —NHSO₂NMe₂,—NHSO₂propyl, —CH₂-morpholine, —CH₂pirolidine, —CH₂NHMe, —SO₂Me,—CH₂SO₂Me, —C≡C—CH₂OMe, —(CH₂)₃OMe, —O(CH₂)₂OMe, —CO₂H, —OH, —NO₂, —CN,—NHSO₂CH₃, and/or phenyl or pyridyl or piperidine, or morpholine whichis preferably unsubstituted.

Het is preferably a 6 to 14 membered ring system and denotes, notwithstanding further substitutions, for example, 2- or 3-furyl, 2- or3-thienyl, 1-, 2- or 3-pyrrolyl, 1-, 2-, 4- or 5-imidazolyl, 1-, 3-, 4-or 5-pyrazolyl, 2-, 4- or 5-oxazolyl, 3-, 4- or 5-isoxazolyl, 2-, 4- or5-thiazolyl, 3-, 4- or 5-isothiazolyl, 2-, 3- or 4-pyridyl, 2-, 4-, 5-or 6-pyrimidinyl, furthermore preferably 1,2,3-triazol-1-, -4- or -5-yl,1,2,4-triazol-1-, -3- or -5-yl, 1- or 5-tetrazolyl, 1,2,3-oxadiazol-4-or -5-yl, 1,2,4-oxadiazol-3- or -5-yl, 1,3,4-thiadiazol-2- or -5-yl,1,2,4-thiadiazol-3- or -5-yl, 1,2,3-thiadiazol-4- or -5-yl, 3- or4-pyridazinyl, pyrazinyl, 1-, 2-, 3-, 4-, 5-, 6- or 7-indolyl,indazolyl, 4- or 5-isoindolyl, 1-, 2-, 4- or 5-benzimidazolyl, 1-, 3-,4-, 5-, 6- or 7-benzopyrazolyl, 2-, 4-, 5-, 6- or 7-benzoxazolyl, 3-,4-, 5-, 6- or 7-benzisoxazolyl, 2-, 4-, 5-, 6- or 7-benzo-thiazolyl, 2-,4-, 5-, 6- or 7-benzisothiazolyl, 4-, 5-, 6- or7-benz-2,1,3-oxadiazolyl, 2-, 3-, 4-, 5-, 6-, 7- or 8-quinolyl, 1-, 3-,4-, 5-, 6-, 7- or 8-isoquinolyl, 3-, 4-, 5-, 6-, 7- or 8-cinnolinyl, 2-,4-, 5-, 6-, 7- or 8-quinazolinyl, 5- or 6-quinoxalinyl, 2-, 3-, 5-, 6-,7- or 8-2H-benzo-1,4-oxazinyl, furthermore preferably1,3-benzodioxol-5-yl, 1,4-benzodioxane-6-yl, 2,1,3-benzothiadiazol-4- or-5-yl or 2,1,3-benzoxadiazol-5-yl.

The heterocyclic radicals may also be partially or fully hydrogenated.

Het can thus also denote, for example, 2,3-dihydro-2-, -3-, -4- or-5-furyl, 2,5-dihydro-2-, -3-, -4- or -5-furyl, tetrahydro-2- or-3-furyl, 1,3-dioxolan-4-yl, tetrahydro-2- or -3-thienyl,2,3-dihydro-1-, -2-, -3-, -4- or -5-pyrrolyl, 2,5-dihydro-1-, -2-, -3-,-4- or -5-pyrrolyl, 1-, 2- or 3-pyrrolidinyl, tetrahydro-1-, -2- or-4-imidazolyl, 2,3-dihydro-1-, -2-, -3-, -4- or -5-pyrazolyl,tetrahydro-1-, -3- or -4-pyrazolyl, 1,4-dihydro-1-, -2-, -3- or-4-pyridyl, 1,2,3,4-tetrahydro-1-, -2-, -3-, -4-, -5- or -6-pyridyl, 1-,2-, 3- or 4-piperidinyl, 2-, 3- or 4-morpholinyl, tetrahydro-2-, -3- or-4-pyranyl, 1,4-dioxaneyl, 1,3-dioxane-2-, -4- or -5-yl, hexahydro-1-,-3- or -4-pyridazinyl, hexahydro-1-, -2-, -4- or -5-pyrimidinyl, 1-, 2-or 3-piperazinyl, 1,2,3,4-tetrahydro-1-, -2-, -3-, -4-, -5-, -6-, -7- or-8-quinolyl, 1,2,3,4-tetrahydro-1-, -2-, -3-, -4-, -5-, -6-, -7- or-8-isoquinolyl, 2-, 3-, 5-, 6-, 7- or8-3,4-dihydro-2H-benzo-1,4-oxazinyl, furthermore preferably2,3-methylenedioxyphenyl, 3,4-methylenedioxyphenyl,2,3-ethylenedioxyphenyl, 3,4-ethylenedioxyphenyl,3,4-(difluoromethylenedioxy)phenyl, 2,3-dihydrobenzofuran-5- or -6-yl,2,3-(2-oxomethylenedioxy)phenyl or also3,4-dihydro-2H-1,5-benzodioxepin-6- or -7-yl, furthermore preferably2,3-dihydrobenzofuranyl or 2,3-dihydro-2-oxofuranyl.

Het very particularly denotes one of the following groups:

wherein X¹, X², and R³ are as defined above.

The compounds of the formula I can have one or more centres of chiralityand can therefore occur in various stereoisomeric forms. The formula Icovers all these forms.

Optionally, the invention relates, in particular, to Formula (I) and itsuse, in which at least one of the said radicals has one of the preferredmeanings indicated above. Some preferred groups of compounds can beexpressed by the following sub-formula Ia to Io, which conform to theformula (I) and in which the radicals not designated in greater detailhave the meaning indicated under the formula I, but in which

-   -   in Ia R^(a) is Ar or Het.    -   in Ib R^(a) is phenyl which is unsubstituted or monosubstituted        or disubstituted, preferably monosubstituted, by F, OCH₃, CH₃,        CF₃, such as, for example, 2′-methoxy-phenyl-,        2′-trifluoromethyl-phenyl-, 2′-chloro-phenyl,        2′,6′-methyl-phenyl-, 2′-alkyl-phenyl-, or pyridyl,    -   in Ic R¹ denotes F,    -   in Id R² denotes H,    -   in Ie Q denotes a single bond in para-position to the        oxadiazole-moiety,        -   S denotes COOH,    -   in If R^(a) denotes heterocyloalkyl preferably bearing at least        an alpha-substituent, such as 2-methyl-piperidin-1-yl,    -   in Ig R¹ is F,        -   R² is H,        -   R^(a) denotes heterocycloalkyl, preferably bearing at least            an alpha-substituent, such as 2-methyl-piperidin-1-yl,        -   R^(b) is trifluoroalkyl,    -   in Ih R¹ is F,        -   R² is H,        -   R^(a) denotes heterocycloalkyl, preferably bearing at least            an alpha-substituent, such as 2-methyl-piperidin-1-yl,        -   R^(b) is nitro,    -   in Ii R¹ is F,        -   R² is H,        -   R^(a) is Ar, preferably bearing at least a 2′ substituent,            such as 2′-methyl-phenyl-, 2′-methoxy-phenyl-,            2′-trifluoromethyl-phenyl-        -   R^(b) is alkyl, nitro, alkoxy,    -   in Ij R¹ is F,        -   R² is H,        -   R^(a) is Het, such as 4-methyl-3-thienyl-        -   R^(b) is alkyl, alkoxy,    -   in Ik R¹ is F,        -   R² is H,        -   R^(a) is Ar, such as 2′-trifluoromethyl-phenyl-,            2′-chloro-phenyl, 2′,6′-methyl-phenyl-, 2′-methyl-phenyl,        -   R^(b) is H,    -   in Il R¹ is F,        -   R² is H,        -   R^(a) is heterocycloalkyl, preferably non-substituted            heterocycloalkyl, such as piperidin-1-yl, morpholinyl        -   R^(b) is nitro, methyl, trifluoromethyl,    -   in Im R¹ is F,        -   R² is H,        -   R^(a) is Ar such as 2′-alkyl-phenyl-,        -   R^(b) is alkyl,    -   in In R¹ is F,        -   R² is H,        -   R^(a) is A or heterocycloalkyl, preferably unsubstituted            such as phenyl, piperidin-1-yl,        -   R^(b) is methyl,    -   In Io R¹ is F        -   R² is H        -   R^(a) is ortho substituted Ar or ortho substituted Het such            as 2-methylphenyl, 2-methylpiperidine, 2-methylmorpholine,            2-methylthienyl.        -   R^(b) is —CH₂OCH₃.

Alternatively, in Formula Ig, Ih, Ii, Ij, Ik, Il, Im, In, and Io, W isCH, Q is a single bond in para position to the oxadiazole moiety and Sis COOH.

Alternatively, in Formula Ig, Ih, Ii, Ij, Ik, Il, Im, In, and Io, QSdenotes

-   -   —COOR³, —CON(R³)(CH₂)_(n)CO₂R³, —CONR³(C₃-C₆cycloalkyl)CO₂R³,        —CH₂N(R³)(CH₂)_(n)CO₂R³, —CH₂NR³(C₃-C₆cycloalkyl)CO₂R³,        —CH₂O(CH₂)_(n)CO₂R³, —CH₂O(C₃-C₆cycloalkyl)CO₂R³,        —O(CH₂)_(n)CO₂R³, —O(C₃-C₆cycloalkyl)CO₂R³ wherein n and R³ are        as above defined.        and pharmaceutically usable derivatives, solvates, salts and        stereoisomers thereof, including mixtures thereof in all ratios.

The compounds of the formula I and also the starting materials for thepreparation thereof are, in addition, prepared by methods known per se,as described in the literature (for example in the standard works, suchas Houben-Weyl, Methoden der organischen Chemie [Methods of OrganicChemistry], Georg-Thieme-Verlag, Stuttgart), under reaction conditionswhich are known and suitable for the said reactions. For all theprotection and deprotection methods, see Philip J. Kocienski, in“Protecting Groups”, Georg Thieme Verlag Stuttgart, New York, 1994 and,Theodora W. Greene and Peter G. M. Wuts in “Protective Groups in OrganicSynthesis”, Wiley Interscience, 3^(rd) Edition 1999.

Use can also be made here of variants which are known per se, but arenot mentioned here in greater detail.

If desired, the starting materials can also be formed in situ so thatthey are not isolated from the reaction mixture, but instead areimmediately converted further into the compounds of the formula I.

The starting compounds for the preparation of compounds of formula I aregenerally known. If they are novel, they can, however, be prepared bymethods known per se.

The reactions are preferably carried out in an inert solvent.

Examples of suitable inert solvents are hydrocarbons, such as hexane,petroleum ether, benzene, toluene or xylene; chlorinated hydrocarbons,such as trichloroethylene, 1,2-dichloroethane, tetrachloromethane,chloroform or DCM; alcohols, such as methanol, ethanol, isopropanol,n-propanol, n-butanol or tert-butanol; ethers, such as diethyl ether,diisopropyl ether, THF (THF) or dioxane; glycol ethers, such as ethyleneglycol monomethyl or monoethyl ether or ethylene glycol dimethyl ether(diglyme); ketones, such as acetone or butanone; amides, such asacetamide, dimethylacetamide or dimethylformamide (DMF); nitriles, suchas acetonitrile; sulfoxides, such as dimethyl sulfoxide (DMSO); carbondisulfide; carboxylic acids, such as formic acid or acetic acid; nitrocompounds, such as nitromethane or nitrobenzene; esters, such as EtOAc,or mixtures of the said solvents.

Pharmaceutical Salts and Other Forms

The said compounds of the formula I can be used in their final non-saltform. On the other hand, the present invention also relates to the useof these compounds in the form of their pharmaceutically acceptablesalts, which can be derived from various organic and inorganic acids andbases by procedures known in the art. Pharmaceutically acceptable saltforms of the compounds of the formula I are for the most part preparedby conventional methods. If the compound of the formula I contains anacidic center, such as a carboxyl group, one of its suitable salts canbe formed by reacting the compound with a suitable base to give thecorresponding base-addition salt. Such bases are, for example, alkalimetal hydroxides, including potassium hydroxide and sodium hydroxide;alkaline earth metal hydroxides, such as magnesium hydroxide and calciumhydroxide; and various organic bases, such as piperidine, diethanolamineand N-methyl-glucamine (meglumine), benzathine, choline, diethanolamine,ethylenediamine, benethamine, diethylamine, piperazine, lysine,L-arginine, ammonia, triethanolamine, betaine, ethanolamine, morpholineand tromethamine. In the case of certain compounds of the formula I,which contain a basic center, acid-addition salts can be formed bytreating these compounds with pharmaceutically acceptable organic andinorganic acids, for example hydrogen halides, such as hydrogen chlorideor hydrogen bromide, other mineral acids and corresponding saltsthereof, such as sulfate, nitrate or phosphate and the like, and alkyl-and monoaryl-sulfonates, such as methanesulfonate, ethanesulfonate,toluenesulfonate and benzene-sulfonate, and other organic acids andcorresponding salts thereof, such as carbonate, acetate,trifluoro-acetate, tartrate, maleate, succinate, citrate, benzoate,salicylate, ascorbate and the like. Accordingly, pharmaceuticallyacceptable acid-addition salts of the compounds of the formula I includethe following: acetate, adipate, alginate, aspartate, benzoate,benzene-sulfonate (besylate), bisulfate, bisulfite, bromide, camphorate,camphor-sulfonate, caprate, caprylate, chloride, chlorobenzoate,citrate, cyclamate, cinnamate, digluconate, dihydrogen-phosphate,dinitrobenzoate, dodecyl-sulfate, ethanesulfonate, formate, glycolate,fumarate, galacterate (from mucic acid), galacturonate, glucoheptanoate,gluco-nate, glutamate, glycerophosphate, hemi-succinate, hemisulfate,heptanoate, hexanoate, hippurate, hydro-chloride, hydrobromide,hydroiodide, 2-hydroxy-ethane-sulfonate, iodide, isethionate,isobutyrate, lactate, lactobionate, malate, maleate, malonate,mandelate, metaphosphate, methanesulfonate, methylbenzoate,mono-hydrogen-phosphate, 2-naphthalenesulfonate, nicotinate, nitrate,oxalate, oleate, palmo-ate, pectinate, persulfate, phenylacetate,3-phenylpropionate, phosphate, phosphonate, phthalate, but this does notrepresent a restriction. Both types of salts may be formed orinterconverted preferably using ion-exchange resin techniques.

Furthermore, the base salts of the compounds of the formula I includealuminum, ammonium, calcium, copper, iron (III), iron(II), lithium,magnesium, manganese(III), manganese(II), potassium, sodium and zincsalts, but this is not intended to represent a restriction. Of theabove-mentioned salts, preference is given to ammonium; the alkali metalsalts sodium and potassium, and the alkaline earth metal salts calciumand magnesium. Salts of the compounds of the formula I which are derivedfrom pharmaceutically acceptable organic non-toxic bases include saltsof primary, secondary and tertiary amines, substituted amines, alsoincluding naturally occurring substituted amines, cyclic amines, andbasic ion exchanger resins for example arginine, betaine, caffeine,chloroprocaine, choline, N,N′-dibenzyl-ethylen-ediamine (benzathine),dicyclohexylamine, diethanol-amine, diethyl-amine,2-diethyl-amino-ethanol, 2-dimethyl-amino-ethanol, ethanolamine,ethylenediamine, N-ethylmorpholine, N-ethyl-piperidine, glucamine,glucosamine, histidine, hydrabamine, isopropyl-amine, lido-caine,lysine, meglumine (N-methyl-D-glucamine), morpholine, piperazine,piperidine, polyamine resins, procaine, purines, theobromine,triethanol-amine, triethylamine, trimethylamine, tripropyl-amine andtris(hydroxy-methyl)-methylamine (tromethamine), but this is notintended to represent a restriction.

Compounds of the formula I of the present invention which contain basicN₂-containing groups can be quaternised using agents such as(C1-C4)-alkyl halides, for example methyl, ethyl, isopropyl andtert-butyl chloride, bromide and iodide; di(C1-C4)alkyl sulfates, forexample dimethyl, diethyl and diamyl sulfate; (C10-C18)alkyl halides,for example decyl, do-decyl, lauryl, myristyl and stearyl chloride,bromide and iodide; and aryl-(C1-C4)alkyl halides, for example benzylchloride and phenethyl bromide. Both water- and oil-soluble compounds ofthe formula I can be prepared using such salts.

The above-mentioned pharmaceutical salts which are preferred includeacetate, trifluoroacetate, besylate, citrate, fumarate, gluconate,hemisuccinate, hippurate, hydrochloride, hydrobromide, isethionate,mandelate, me-glumine, nitrate, oleate, phosphonate, pivalate, sodiumphosphate, stearate, sulfate, sulfosalicylate, tartrate, thiomalate,tosylate and tro-meth-amine, but this is not intended to represent arestriction.

The acid-addition salts of basic compounds of the formula I are preparedby bringing the free base form into contact with a sufficient amount ofthe desired acid, causing the formation of the salt in a conventionalmanner. The free base can be regenerated by bringing the salt form intocontact with a base and isolating the free base in a conventionalmanner. The free base forms differ in a certain respect from thecorresponding salt forms thereof with respect to certain physicalproperties, such as solubility in polar solvents; for the purposes ofthe invention, however, the salts other-wise correspond to therespective free base forms thereof.

As mentioned, the pharmaceutically acceptable base-addition salts of thecompounds of the formula I are formed with metals or amines, such asalkali metals and alkaline earth metals or organic amines. Preferredmetals are sodium, potassium, magnesium and calcium. Preferred organicamines are N,N′-dibenzylethylenediamine, chloroprocaine, choline,diethanol-amine, ethylenediamine, N-methyl-D-glucamine and procaine.

The base-addition salts of acidic compounds of the formula I areprepared by bringing the free acid form into contact with a sufficientamount of the desired base, causing the formation of the salt in aconventional manner. The free acid can be regenerated by bringing thesalt form into contact with an acid and isolating the free acid in aconventional manner. The free acid forms differ in a certain respectfrom the corresponding salt forms thereof with respect to certainphysical properties, such as solubility in polar solvents; for thepurposes of the invention, however, the salts other-wise correspond tothe respective free acid forms thereof.

If a compound of the formula I contains more than one group which iscapable of forming pharmaceutically acceptable salts of this type, theformula I also encompasses multiple salts. Typical multiple salt formsinclude, for example, bitartrate, diacetate, difumarate, dimeglumine,di-phosphate, disodium and trihydrochloride, but this is not intended torepresent a restriction.

With regard to that stated above, it can be seen that the term“pharmaceutically acceptable salt” in the present connection is taken tomean an active ingredient which comprises a compound of the formula I inthe form of one of its salts, in particular if this salt form impartsimproved pharmacokinetic properties on the active ingredient comparedwith the free form of the active ingredient or any other salt form ofthe active ingredient used earlier. The pharmaceutically acceptable saltform of the active ingredient can also provide this active ingredientfor the first time with a desired pharmacokinetic property which it didnot have earlier and can even have a positive influence on thepharmacodynamics of this active ingredient with respect to itstherapeutic efficacy in the body.

Owing to their molecular structure, the compounds of the formula I canbe chiral and can accordingly occur in various enantiomeric forms. Theycan therefore exist in racemic or in optically active form.

Since the pharmaceutical activity of the racemates or stereoisomers ofthe compounds according to the invention may differ, it may be desirableto use the enantiomers. In these cases, the end product or even theIntermediates can be separated into enantiomeric compounds by chemicalor physical measures known to the person skilled in the art or evenemployed as such in the synthesis.

In the case of racemic amines, diastereomers are formed from the mixtureby reaction with an optically active resolving agent. Examples ofsuitable resolving agents are optically active acids, such as the R andS forms of tartaric acid, diacetyltartaric acid, dibenzoyltartaric acid,mandelic acid, malic acid, lactic acid, suitable N-protected amino acids(for example N-benzoylproline or N-benzenesulfonylproline), or thevarious optically active camphorsulfonic acids. Also advantageous ischromatographic enantiomer resolution with the aid of an opticallyactive resolving agent (for example dinitrobenzoylphenylglycine,cellulose triacetate or other derivatives of carbohydrates or chirallyderivatised methacrylate polymers immobilised on silica gel). Suitableeluents for this purpose are aqueous or alcoholic solvent mixtures, suchas, for example, hexane/isopropanol/acetonitrile, for example in theratio 82:15:3.

The invention furthermore relates to the use of compounds of formula I,in combination with at least one further medicament active ingredient,preferably medicaments used in the treatment of multiple sclerosis suchas cladribine or another co-agent, such as interferon, e.g. pegylated ornon-pegylated interferons, preferably interferon beta and/or withcompounds improving vascular function. These further medicaments, suchas interferon beta, may be administered concomitantly or sequentially,e.g. by subcutaneous, intramuscular or oral routes. These compositionscan be used as medicaments in human and veterinary medicine.

Pharmaceutical formulations can be administered in the form of dosageunits, which comprise a predetermined amount of active ingredient perdosage unit. Such a unit can comprise, for example, 0.5 mg to 1 g,preferably 1 mg to 700 mg, particularly preferably 5 mg to 100 mg, of acompound according to the invention, depending on the disease conditiontreated, the method of administration and the age, weight and conditionof the patient, or pharmaceutical formulations can be administered inthe form of dosage units which comprise a predetermined amount of activeingredient per dosage unit. Preferred dosage unit formulations are thosewhich comprise a daily dose or part-dose, as indicated above, or acorresponding fraction thereof of an active ingredient. Furthermore,pharmaceutical formulations of this type can be prepared using aprocess, which is generally known in the pharmaceutical art.

Pharmaceutical formulations can be adapted for administration via anydesired suitable method, for example by oral (including buccal orsublingual), rectal, nasal, topical (including buccal, sublingual ortransdermal), vaginal or parenteral (including subcutaneous,intramuscular, intravenous or intradermal) methods. Such formulationscan be prepared using all processes known in the pharmaceutical art by,for example, combining the active ingredient with the excipient(s) oradjuvant(s).

Pharmaceutical formulations adapted for oral administration can beadministered as separate units, such as, for example, capsules ortablets; powders or granules; solutions or suspensions in aqueous ornon-aqueous liquids; edible foams or foam foods; or oil-in-water liquidemulsions or water-in-oil liquid emulsions.

Thus, for example, in the case of oral administration in the form of atablet or capsule, the active-ingredient component can be combined withan oral, non-toxic and pharmaceutically acceptable inert excipient, suchas, for example, ethanol, glycerol, water and the like. Powders areprepared by comminuting the compound to a suitable fine size and mixingit with a pharmaceutical excipient comminuted in a similar manner, suchas, for example, an edible carbohydrate, such as, for example, starch ormannitol. A flavour, preservative, dispersant and dye may likewise bepresent.

Capsules are produced by preparing a powder mixture as described aboveand filling shaped gelatine shells therewith. Glidants and lubricants,such as, for example, highly disperse silicic acid, talc, magnesiumstearate, calcium stearate or polyethylene glycol in solid form, can beadded to the powder mixture before the filling operation. A disintegrantor solubiliser, such as, for example, agar-agar, calcium carbonate orsodium carbonate, may likewise be added in order to improve theavailability of the medica-ment after the capsule has been taken.

In addition, if desired or necessary, suitable binders, lubricants anddisintegrants as well as dyes can likewise be incorporated into themixture. Suitable binders include starch, gelatine, natural sugars, suchas, for example, glucose or beta-lactose, sweeteners made from maize,natural and synthetic rubber, such as, for example, acacia, tragacanthor sodium alginate, carboxymethylcellulose, polyethylene glycol, waxes,and the like. The lubricants used in these dosage forms include sodiumoleate, sodium stearate, magnesium stearate, sodium benzoate, sodiumacetate, sodium chloride and the like. The disintegrants include,without being restricted thereto, starch, methylcellulose, agar,bentonite, xanthan gum and the like. The tablets are formulated by, forexample, preparing a powder mixture, granulating or dry-pressing themixture, adding a lubricant and a disintegrant and pressing the entiremixture to give tablets. A powder mixture is prepared by mixing thecompound comminuted in a suitable manner with a diluent or a base, asdescribed above, and optionally with a binder, such as, for example,carboxymethylcellulose, an alginate, gelatine or polyvinyl-pyrrolidone,a dissolution retardant, such as, for example, paraffin, an absorptionaccelerator, such as, for example, a quaternary salt, and/or anabsorbent, such as, for example, bentonite, kaolin or dicalciumphosphate. The powder mixture can be granulated by wetting it with abinder, such as, for example, syrup, starch paste, acadia mucilage orsolutions of cellulose or polymer materials and pressing it through asieve. As an alternative to granulation, the powder mixture can be runthrough a tableting machine, giving lumps of non-uniform shape which arebroken up to form granules. The granules can be lubricated by additionof stearic acid, a stearate salt, talc or mineral oil in order toprevent sticking to the tablet casting moulds. The lubricated mixture isthen pressed to give tablets. The active ingredients can also becombined with a free-flowing inert excipient and then pressed directlyto give tablets without carrying out the granulation or dry-pressingsteps. A transparent or opaque protective layer consisting of a shellacsealing layer, a layer of sugar or polymer material and a gloss layer ofwax may be present. Dyes can be added to these coatings in order to beable to differentiate between different dosage units.

Oral liquids, such as, for example, solution, syrups and elixirs, can beprepared in the form of dosage units so that a given quantity comprisesa pre-specified amount of the compounds. Syrups can be prepared bydissolving the compounds in an aqueous solution with a suitable flavour,while elixirs are prepared using a non-toxic alcoholic vehicle.Suspensions can be formulated by dispersion of the compounds in anon-toxic vehicle. Solubilisers and emulsifiers, such as, for example,ethoxylated isostearyl alcohols and polyoxyethylene sorbitol ethers,preservatives, flavour additives, such as, for example, peppermint oilor natural sweeteners or saccharin, or other artificial sweeteners andthe like, can likewise be added.

The dosage unit formulations for oral administration can, if desired, beencapsulated in microcapsules. The formulation can also be prepared insuch a way that the release is extended or retarded, such as, forexample, by coating or embedding of particulate material in polymers,wax and the like.

The compounds of the formula I and salts, solvates and physiologicallyfunctional derivatives thereof and the other active ingredients can alsobe administered in the form of liposome delivery systems, such as, forexample, small unilamellar vesicles, large unilamellar vesicles andmultilamellar vesicles. Liposomes can be formed from variousphospholipids, such as, for example, cholesterol, stearylamine orphosphatidylcholines.

The compounds of the formula I and the salts, solvates andphysiologically functional derivatives thereof and the other activeingredients can also be delivered using monoclonal antibodies asindividual carriers to which the compound molecules are coupled. Thecompounds can also be coupled to soluble polymers as targeted medicamentcarriers. Such polymers may encompass polyvinylpyrrolidone, pyrancopolymer, polyhydroxypropyl-methacrylamidophenol,polyhydroxyethylaspartamidophenol or polyethylene oxide polylysine,substituted by palmitoyl radicals. The compounds may furthermore becoupled to a class of biodegradable polymers which are suitable forachieving controlled release of a medicament, for example polylacticacid, poly-epsilon-caprolactone, polyhydroxybutyric acid,poly-orthoesters, polyacetals, polydihydroxypyrans, polycyanoacrylatesand crosslinked or amphipathic block copolymers of hydrogels.

Pharmaceutical formulations adapted for transdermal administration canbe administered as independent plasters for extended, close contact withthe epidermis of the recipient. Thus, for example, the active ingredientcan be delivered from the plaster by iontophoresis, as described ingeneral terms in Pharmaceutical Research, 3(6), 318 (1986).

Pharmaceutical compounds adapted for topical administration can beformulated as ointments, creams, suspensions, lotions, powders,solutions, pastes, gels, sprays, aerosols or oils.

For the treatment of the eye or other external tissue, for example mouthand skin, the formulations are preferably applied as topical ointment orcream. In the case of formulation to give an ointment, the activeingredient can be employed either with a paraffinic or a water-misciblecream base. Alternatively, the active ingredient can be formulated togive a cream with an oil-in-water cream base or a water-in-oil base.

Pharmaceutical formulations adapted for topical application to the eyeinclude eye drops, in which the active ingredient is dissolved orsuspended in a suitable carrier, in particular an aqueous solvent.

Pharmaceutical formulations adapted for topical application in the mouthencompass lozenges, pastilles and mouthwashes.

Pharmaceutical formulations adapted for rectal administration can beadministered in the form of suppositories or enemas.

Pharmaceutical formulations adapted for nasal administration in whichthe carrier substance is a solid comprise a coarse powder having aparticle size, for example, in the range 20-500 microns, which isadministered in the manner in which snuff is taken, i.e. by rapidinhalation via the nasal passages from a container containing the powderheld close to the nose. Suitable formulations for administration asnasal spray or nose drops with a liquid as carrier substance encompassactive-ingredient solutions in water or oil.

Pharmaceutical formulations adapted for administration by inhalationencompass finely particulate dusts or mists, which can be generated byvarious types of pressurised dispensers with aerosols, nebulisers orinsuf-flators.

Pharmaceutical formulations adapted for vaginal administration can beadministered as pessaries, tampons, creams, gels, pastes, foams or sprayformulations.

Pharmaceutical formulations adapted for parenteral administrationinclude aqueous and non-aqueous sterile injection solutions comprisingantioxidants, buffers, bacteriostatics and solutes, by means of whichthe formulation is rendered isotonic with the blood of the recipient tobe treated; and aqueous and non-aqueous sterile suspensions, which maycomprise suspension media and thickeners. The formulations can beadministered in single-dose or multidose containers, for example sealedampoules and vials, and stored in freeze-dried (lyophilised) state, sothat only the addition of the sterile carrier liquid, for example waterfor injection purposes, immediately before use is necessary.

Injection solutions and suspensions prepared in accordance with therecipe can be prepared from sterile powders, granules and tablets.

It goes without saying that, in addition to the above particularlymentioned constituents, the formulations may also comprise other agentsusual in the art with respect to the particular type of formulation;thus, for example, formulations which are suitable for oraladministration may comprise flavours.

A therapeutically effective amount of a compound of the formula I and ofthe other active ingredient depends on a number of factors, including,for example, the age and weight of the animal, the precise diseasecondition which requires treatment, and its severity, the nature of theformulation and the method of administration, and is ultimatelydetermined by the treating doctor or vet. However, an effective amountof a compound is generally in the range from 0.1 to 100 mg/kg of bodyweight of the recipient (mammal) per day and particularly typically inthe range from 1 to 10 mg/kg of body weight per day. Thus, the actualamount per day for an adult mammal weighing 70 kg is usually between 70and 700 mg, where this amount can be administered as an individual doseper day or usually in a series of part-doses (such as, for example, two,three, four, five or six) per day, so that the total daily dose is thesame. An effective amount of a salt or solvate or of a physiologicallyfunctional derivative thereof can be determined as the fraction of theeffective amount of the compound per se.

The present invention furthermore relates to a method for treating asubject suffering from a sphingosine 1-phosphate associated disorder,comprising administering to said subject an effective amount of acompounds of formula I. The present invention preferably relates to amethod, wherein the sphingosine 1-phosphate-1 associated disorder is anautoimmune disorder or condition associated with an overactive immuneresponse.

The present invention furthermore relates to a method of treating asubject suffering from an immunoregulatory abnormality, comprisingadministering to said subject a compounds of formula I in an amount thatis effective for treating said immunoregulatory abnormality. The presentinvention preferably relates to a method wherein the immunoregulatoryabnormality is an autoimmune or chronic inflammatory disease selectedfrom the group consisting of: amyotrophic lateral sclerosis (ALS),systemic lupus erythematosus, chronic rheumatoid arthritis, type Idiabetes mellitus, inflammatory bowel disease, biliary cirrhosis,uveitis, multiple sclerosis, Crohn's disease, ulcerative colitis,bullous pemphigoid, sarcoidosis, psoriasis, autoimmune myositis,Wegener's granulomatosis, ichthyosis, Graves' ophthalmopathy and asthma.The present invention furthermore relates to a method wherein theimmunoregulatory abnormality is bone marrow or organ transplantrejection or graft-versus-host disease. The present inventionfurthermore relates to a method wherein the immunoregulatory abnormalityis selected from the group consisting of: transplantation of organs ortissue, graft-versus-host diseases brought about by transplantation,autoimmune syndromes including rheumatoid arthritis, systemic lupuserythematosus, Hashimoto's thyroiditis, multiple sclerosis, myastheniagravis, type I diabetes, uveitis, posterior uveitis, allergicencephalomyelitis, glomerulonephritis, post-infectious autoimmunediseases including rheumatic fever and post-infectiousglomerulonephritis, inflammatory and hyperproliferative skin diseases,psoriasis, atopic dermatitis, contact dermatitis, eczematous dermatitis,seborrhoeic dermatitis, lichen planus, pemphigus, bullous pemphigoid,epidermolysis bullosa, urticaria, angioedemas, vasculitis, erythema,cutaneous eosinophilia, lupus erythematosus, acne, alopecia greata,keratoconjunctivitis, vernal conjunctivitis, uveitis associated withBehcet's disease, keratitis, herpetic keratitis, conical cornea,dystrophia epithelialis corneae, corneal leukoma, ocular pemphigus,Mooren's ulcer, scleritis, Graves' opthalmopathy, Vogt-Koyanagi-Haradasyndrome, sarcoidosis, pollen allergies, reversible obstructive airwaydisease, bronchial asthma, allergic asthma, intrinsic asthma, extrinsicasthma, dust asthma, chronic or inveterate asthma, late asthma andairway hyper-responsiveness, bronchitis, gastric ulcers, vascular damagecaused by ischemic diseases and thrombosis, ischemic bowel diseases,inflammatory bowel diseases, necrotizing enterocolitis, intestinallesions associated with thermal burns, coeliac diseases, proctitis,eosinophilic gastroenteritis, mastocytosis, Crohn's disease, ulcerativecolitis, migraine, rhinitis, eczema, interstitial nephritis,Goodpasture's syndrome, hemolytic-uremic syndrome, diabetic nephropathy,multiple myositis, Guillain-Barre syndrome, Meniere's disease,polyneuritis, multiple neuritis, mononeuritis, radiculopathy,hyperthyroidism, Basedow's disease, pure red cell aplasia, aplasticanemia, hypoplastic anemia, idiopathic thrombocytopenic purpura,autoimmune hemolytic anemia, agranulocytosis, pernicious anemia,megaloblastic anemia, anerythroplasia, osteoporosis, sarcoidosis,fibroid lung, idiopathic interstitial pneumonia, dermatomyositis,leukoderma vulgaris, ichthyosis vulgaris, photoallergic sensitivity,cutaneous T cell lymphoma, chronic lymphocytic leukemia,arteriosclerosis, atherosclerosis, aortitis syndrome, polyarteritisnodosa, myocardosis, scleroderma, Wegener's granuloma, Sjogren'ssyndrome, adiposis, eosinophilic fascitis, lesions of gingiva,periodontium, alveolar bone, substantia ossea dentis,glomerulonephritis, male pattern alopecia or alopecia senilis bypreventing epilation or providing hair germination and/or promoting hairgeneration and hair growth, muscular dystrophy, pyoderma and Sezary'ssyndrome, Addison's disease, ischemia-reperfusion injury of organs whichoccurs upon preservation, transplantation or ischemic disease,endotoxin-shock, pseudomembranous colitis, colitis caused by drug orradiation, ischemic acute renal insufficiency, chronic renalinsufficiency, toxinosis caused by lung-oxygen or drugs, lung cancer,pulmonary emphysema, cataracta, siderosis, retinitis pigmentosa, senilemacular degeneration, vitreal scarring, corneal alkali burn, dermatitiserythema multiforme, linear IgA ballous dermatitis and cementdermatitis, gingivitis, periodontitis, sepsis, pancreatitis, diseasescaused by environmental pollution, aging, carcinogenesis, metastasis ofcarcinoma and hypobaropathy, disease caused by histamine orleukotriene-C₄ release, Behcet's disease, autoimmune hepatitis, primarybiliary cirrhosis, sclerosing cholangitis, partial liver resection,acute liver necrosis, necrosis caused by toxin, viral hepatitis, shock,or anoxia, B-virus hepatitis, non-A/non-B hepatitis, cirrhosis,alcoholic cirrhosis, hepatic failure, fulminant hepatic failure,late-onset hepatic failure, “acute-on-chronic” liver failure,augmentation of chemotherapeutic effect, cytomegalovirus infection, HCMVinfection, AIDS, cancer, senile dementia, trauma, and chronic bacterialinfection.

Preferred compounds of formula (I) exhibit a EC50 in GTPγS for thebinding to the S1P₁ receptor of less than about 10 μM, preferably lessthan about 5 μM, more preferably less than about 1 μM and even morepreferred less than about 0.1 μM. Most preferably, compounds of Formula(I) exhibit a EC50 for the binding of S1P1 less than 0.01 μM.

Preferred compounds of Formula (I) exhibit a selectivity on S1P1receptor over the S1P3 receptor of a magnitude of more than about 20.More preferably, compounds of formula (I) are 50 fold selective for S1P1compare to S1P3, more preferably, 100 fold.

The compounds of invention have been named according the standards usedin the program “ACD/Name Batch” from Advanced Chemistry DevelopmentInc., ACD/Labs (7.00 Release). Product version: 7.10, build: 15 Sep.2003.

In the following the present invention shall be illustrated by means ofsome examples, which are not construed to be viewed as limiting thescope of the invention.

EXAMPLES

Compounds of the present invention are synthesized according to theprotocols described in the patent application PCT/EP2008/063180

The oxadiazole compounds according to formula (I) can be prepared fromreadily available starting materials by several synthetic approaches,using both solution-phase and solid-phase chemistry protocols or mixedsolution and solid phase protocols. Examples of synthetic pathways aredescribed below in the examples.

The commercially available starting materials used in the followingexperimental description were purchased from Aldrich or Fluke unlessotherwise reported.

The HPLC, NMR and MS data provided in the examples described below areobtained as followed:

HPLC data:

Method A: HPLC columns: Xbridge™ C8 column 50 mm×4.6 mm at a flow of 2mL/min; 8 min gradient from 0.1% TFA in H₂O to 0.07% TFA in ACN.

Method B: HPLC columns: ATLANTIS C18 75×4.6 mm 5 U at a flow of 1mL/min; A—0.1% HCOOH B-ACN.

Method C: HPLC columns: 018 BDS, 50×4.6 mm, SC\307 at a flow of 0.8mL/min; A—0.1% TFA, B—ACN: Flow—0.8 mL/min.

Method D: HPLC columns: ATLANTIS C18 75×4.6 mm, 5 U at a flow of 0.8ml/min; A—0.1% TFA, B—ACN

Method: E: HPLC columns: ATLANTIS C18 75×4.6 mm, 5 U at a flow of 0.8ml/min; A—10 mM NH₄OAC, B—ACN

Method F: HPLC columns: Phenomenex Luna 5 μm C18 (2), 100×4.6 mm (plusguard cartridge) at a flow of 2 ml/min; 3.5 min gradient from 95:5([0.1% (V/V) formic acid in H₂O]: [0.1% (V/V) formic acid in MeCN]) to5:95% ([0.1% (V/V) formic acid in H₂O]: [0.1% (V/V) formic acid inMeCN]) then held for 2 minutes at 5:95 ([0.1% (V/V) formic acid in H₂O]:[0.1% (V/V) formic acid in MeCN]).

Method G: HPLC columns: Waters Xterra MS 5 μm C18, 100×4.6 mm (plusguard cartridge) at a flow of 2 ml/min; 3.5 min gradient from 95:5 ([10mM ammonium bicarbonate in H₂O]: MeCN) to 5:95 ([10 mM ammoniumbicarbonate in H₂O]: MeCN) then held for 1.5 minutes at 5:95 ([10 mMammonium bicarbonate in H₂O]: MeCN).

Method H: HPLC columns: Waters Sunfire 5 μm C18, 150×4.6 mm (plus guardcartridge) at a flow of 1 ml/min; 30 min gradient from 95:5 ([0.1% (V/V)formic acid in H₂O]: [0.1% (V/V) formic acid in MeOH]) to 0.1% (V/V)formic acid in MeOH then held for 5 minutes at 0.1% (V/V) formic acid inMeOH.

Method I: Gradient of Method H applied to HPLC columns: Supelco,Ascentis® Express 018 or Hichrom Halo C18, 2.7 μm C18, 100×4.6 mm.

Method J: HPLC columns: Waters Xterra 5 μm C18 (2), 250×4.6 mm (plusguard cartridge) at a flow of 1 ml/min; 19.5 min gradient from 95:5 ([10mM ammonium bicarbonate in H₂O]: MeCN) to MeCN then held for 4 minutesat MeCN.

Method K: HPLC columns: Waters Xbridge 5 μm C18, 150×4.6 mm (plus guardcartridge) at a flow of 1 ml/min; 22 min gradient from 95:5 ([10 mMammonium bicarbonate in H₂O]: MeOH) to MeOH then held for 4 minutes atMeOH.

UV detection (maxplot) for all methods.

Mass Spectrum:

Method A: LC/MS Waters ZMD (ESI); GC/MS: GC Agilent 6890N & MS Agilent5973.

Method B: UPLC/MS: Waters Acquity, column Waters Acquity UPLC BEH C181.7 m 2.1×50 mm, conditions: solvent A (10 mM ammonium acetate inwater+5% ACN), solvent B (ACN), gradient 5% B to 100% B over 3 min, UVdetection (PDA, 230-400 nm) and MS detection (SQ detector, positive andnegative ESI modes, cone voltage 30 V).

¹H-NMR data:

Bruker DPX-300 MHz unless otherwise reported.

Preparative HPLC Purifications:

Preparative HPLC purifications were performed with HPLC waters Prep LC4000 System equipped with Columns® PrepMS C18 10m, 50×300 mm, unlessotherwise reported. All HPLC purifications were performed with agradient of ACN/H₂O or ACN/H₂O/TFA (0.1%).

Mass Directed Autoprep Purifications:

Preparative HPLC purifications are performed with a mass directedautopurification Fractionlynx from Waters equipped with a Sunfire PrepC18 OBD column 19×100 mm 5 m, unless otherwise reported. All HPLCpurifications were performed with a gradient of ACN/H₂O or ACN/H₂O/HCOOH(0.1%).

The microwave chemistry was performed on a single mode microwave reactorEmrys™ Optimiser from Personal Chemistry.

General Procedures:

Procedure 1: Amidoxime Moiety Formation

To a solution of nitrile derivative (1 eq) in EtOH (1-5 mL/mmol ofnitrile derivative) was added a 50% aqueous solution of NH₂OH (5 eq).The resulting mixture was stirred at a temperature ranging from RT to80° C. for 1 to 72 hours. In case a precipitation of the expectedcompound was observed, the precipitate was filtered off and washed withan adequate solvent, such as EtOH, iPrOH or water, and then dried underreduced pressure to give the expected amidoxime derivative. In all othercases, the reaction mixture was concentrated under reduced pressure,diluted with an adequate solvent, such as water or iPrOH, until theprecipitation of the expected compound was observed. The precipitate wasfiltered off and washed with an adequate solvent, such as iPrOH orwater, and then dried under reduced pressure to give the expectedamidoxime derivative. When no precipitation occurred, the concentratedmixture was dissolved in EtOAc and water, the organic layer was washedwith water (twice) and brine (twice), then dried over MgSO₄, filteredand concentrated under vacuum to give the expected amidoxime derivative.

Procedure 2: Amidoxime Moiety Formation

To a solution of nitrile derivative (1 eq) in EtOH (1-5 mL/mmol ofnitrile derivative) was added NH₂OH.HCl (1.1 eq) and Et₃N (1.2 eq). Theresulting mixture was stirred at a temperature ranging from RT to 80° C.for 1 to 72 hours. In case of precipitation of the expected compound,the precipitate was filtered off and washed with an adequate solvent,such as EtOH, iPrOH or water, and then dried under reduced pressure togive the expected amidoxime derivative. In all other cases, the reactionmixture was concentrated under reduced pressure, diluted with anadequate solvent, such as water or iPrOH, until precipitation. Theprecipitate was filtered off and washed with an adequate solvent, suchas iPrOH or water, and then dried under reduced pressure to give theexpected amidoxime derivative. When no precipitation occurred, theconcentrated mixture was dissolved in EtOAc and water, the organic layerwas washed with water (twice) and brine (twice), then dried over MgSO₄,filtered and concentrated under vacuum to give the expected amidoximederivative.

Procedure 3: Oxadiazole Ring Formation

DIEA (2.0 to 2.2 eq) and HATU (1.0 to 1.1 eq) were added into a solutionof the carboxylic acid derivative (1 eq) in anhydrous DMF (4 mL/mmol ofcarboxylic acid derivative) cooled at 0° C. The resulting mixture wasstirred at 0° C. for a period of 5 to 30 minutes. Then the amidoximederivative (1.0 to 1.2 eq) was added neat or as a DMF solution. Theresulting mixture was stirred at 0° C. or RT for a period of 30 minutesto 18 hours. The reaction mixture was diluted with an adequate solvent,such as Et₂O, MTBE or EtOAc, and then washed with water and brine. Theaqueous layers were extracted once. The organic layers were combined,dried (MgSO₄ or Na₂SO₄) and the solvents were removed under reducedpressure. The residue was either taken up with toluene (6 mL/mmol ofcarboxylic acid derivative) and pyridine (2 mL/mmol of carboxylic acidderivative) or with ACN (8.5 mL/mmol of carboxylic acid derivative). Theresulting mixture was heated at a temperature between 80° C. to refluxfor a period of 12 to 72 hours. The reaction mixture was diluted with anadequate solvent, such as Et₂O, MTBE or EtOAc, and then washed withwater and brine. The aqueous layers were extracted once. The organiclayers were combined, dried (MgSO₄ or Na₂SO₄) and the solvents wereevaporated under reduced pressure. Purification by flash chromatographyor precipitation gave the expected oxadiazole derivative.

Procedure 4: Oxadiazole Ring Formation

DIEA (2.0 to 2.2 eq) and HATU (1.0 to 1.1 eq) were added into a solutionof the carboxylic acid derivative (1 eq) in anhydrous DMF (4 mL/mmol ofcarboxylic acid derivative) cooled at 0° C. The resulting mixture wasstirred at 0° C. for a period of 5 to 30 minutes. Then the amidoximederivative (1.0 to 1.2 eq) was added neat or as a DMF solution. Theresulting mixture was stirred at 0° C. or RT for a period of 30 minutesto 18 hours. The reaction mixture was diluted with an adequate solvent,such as Et₂O, MTBE or EtOAc, and then washed with water and brine. Theaqueous layers were extracted once. The organic layers were combined,dried (MgSO₄ or Na₂SO₄) and the solvents were removed under reducedpressure. The residue was taken up with ACN (8.5 mL/mmol of carboxylicacid derivative). The resulting mixture was heated at 150° C. for 30 minunder MW irradiation. The reaction mixture was diluted with an adequatesolvent, such as Et₂O, MTBE or EtOAc, and then washed with water andbrine. The aqueous layers were extracted once. The organic layers werecombined, dried (MgSO₄ or Na₂SO₄) and the solvents were evaporated underreduced pressure. Purification by flash chromatography or precipitationgave the expected oxadiazole derivative.

Procedure 5: Oxadiazole Ring Formation

To a suspension of carboxylic acid derivative (1 eq) in dry toluene (3.5mL/mmol of carboxylic acid) at RT under N₂ was added oxalyl chloride(1.5 eq) and DMF (0.02 eq). The reaction mixture was stirred at RT for 2hours and then concentrated under vacuum. The resulting acid chloridewas then dissolved in dry toluene (2.5 mL/mmol of carboxylic acid) andadded dropwise at RT under N₂ to a solution of amidoxime derivative (1eq) in pyridine (1.3 mL/mmol of amidoxime) and toluene (1.3 mL/mmol ofamidoxime). The reaction mixture was stirred at RT for 1 hour and thenrefluxed overnight. The reaction mixture was cooled to RT andconcentrated under vacuum. The resulting oil was taken in EtOAc, washedwith water, saturated solution of NaHCO₃ (twice), saturated solution ofNaCl, then dried over MgSO₄, filtered and concentrated under vacuum.Purification by flash chromatography or precipitation gave the expectedoxadiazole derivative.

Procedure 7: Oxadiazole Ring Formation

In a microwave vessel was added carboxylic acid derivative (1 eq),amidoxime derivative (1.1 eq),1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (1.1 eq),THF (2.7 mL /mmol of carboxylic acid) and CH₃CN (2.7 mL/mmol ofcarboxylic acid). The reaction mixture was stirred at RT for 2 hours andwas then heated at 150° C. for 30 minutes under microwave irradiation.The reaction mixture was passed through SPE NH₂ and SPE SCX rinsed withACN and the solvents were evaporated. Purification by flashchromatography or precipitation gave the expected oxadiazole derivative.

Procedure 6: Oxadiazole Ring Formation

NMM (3 to 5 eq) and isobutyl chloroformate (1.0 to 1.1 eq) were addedinto a solution of the carboxylic acid derivative (1.0 eq) in a suitablesolvent, such as dioxane or isopropyl acetate and stirred at atemperature comprised between 0° C. and RT for 10 minutes to a fewhours. Then the amidoxime derivative (1.0 to 1.2 eq) was added in oneportion and the reaction mixture was stirred at a temperature comprisedbetween 0° C. and RT for 20 min to a few hours, and then heated at atemperature comprised between 80° C. to reflux for a period of 12 to 24hours. The reaction mixture was diluted with an adequate solvent, suchas Et₂O, MTBE or EtOAc, and then washed with water and brine. Theaqueous layers were extracted once. The organic layers were combined,dried (MgSO₄ or Na₂SO₄) and the solvents were evaporated under reducedpressure. Purification by flash chromatography or precipitation gave theexpected oxadiazole derivative.

Procedure 7: Oxadiazole Ring Formation

In a microwave vessel was added carboxylic acid derivative (1 eq),amidoxime derivative (1.1 eq),1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (1.1 eq),THF (2.7 mL/mmol of carboxylic acid) and CH₃CN (2.7 mL/mmol ofcarboxylic acid). The reaction mixture was stirred at RT for 2 hours andwas then heated at 150° C. for 30 minutes under microwave irradiation.The reaction mixture was passed through SPE NH₂ and SPE SCX rinsed withACN and the solvents were evaporated. Purification by flashchromatography or precipitation gave the expected oxadiazole derivative.

Procedure 8: Tert-Butyl Ester Hydrolysis

To tert-butyl ester derivative (1 eq) was added hydrochloric acid indioxane (4N, 20-50 eq) and the reaction mixture was stirred at RT for 1hour to 24 hours. The solution was then evaporated to dryness and theresidue was purified by precipitation from a solvent such as CH₃CN, DCM,MTBE or Et₂O to afford the title compound.

Procedure 9: Methyl or Ethyl Ester Hydrolysis

To the methyl or ethyl ester derivative (1 eq) in a solution of MeOH orEtOH (5 mL/mmol of ester derivative) or MeOH/THF 1:1 or EtOH/THF 1:1 (5mL/mmol of ester derivative) was added sodium hydroxide (5M, 5 eq) andthe reaction mixture was stirred at RT for 1 hour to 24 hours. Thesolution was evaporated to dryness. The residue was taken up with EtOAcand washed with a 1N aqueous solution of HCl and brine. The organiclayer was dried (MgSO₄) and concentrated under vacuum to afford thetitle compound.

Procedure 10: Substitution of Benzyl Bromides with Primary and SecondaryAmines

To a solution of bromobenzyl derivative (1 eq) in CH₃CN or DMF (1.5-3mL/mmol of bromobenzyl derivative) was added K₂CO₃ or NaHCO₃ (2-3 eq)and a primary or secondary amine (1.2 eq). The reaction mixture wasstirred at RT or at 60° C. overnight. Solvent was removed under vacuumand the resulting mixture was diluted with water, extracted with EtOAc,washed with a saturated solution of NaCl, dried over MgSO₄ andconcentrated under vacuum. Purification by flash chromatography orrecrystallization afforded the expected benzyl amine derivative.

Procedure 11: Suzuki Cross-Coupling Reaction

A mixture of the aryl bromide (1 eq), the boronic acid or esterderivative (1.2 to 2.0 eq), cesium fluoride (3 eq) andbis(triphenylphosphine)palladium(II) chloride (0.01 to 0.05 eq) wasprepared in dioxane (10 mL/g of aryl bromide) and water (4 mL/g of arylbromide) under nitrogen atmosphere. The resulting mixture was heated ata temperature comprised between 80° C. to reflux for 1 to 15 hours. Thereaction mixture was diluted with MTBE (40 mL/g of aryl bromide) and theaqueous layer was removed. The organic layer was dried (MgSO₄ or Na₂SO₄)and concentrated under vacuum. The residue was purified by flashchromatography.

Procedure 12: Aromatic Cyanation

A mixture of aryl halide (1 eq), sodium cyanide (1.5 eq) andtetrabutylammonium bromide (1.5 eq) was dissolved in DMA (2 mL/mmol ofaryl halide) and heated at 120° C. for 1 to 3 hours under microwaveirradiation. The resulting mixture was diluted with EtOAc, washed with asaturated solution of NaCl, dried over MgSO₄ and concentrated undervacuum. Purification by flash chromatography or recrystallizationafforded the expected aryl cyanide derivative.

Procedure 13: Benzylic Bromation

To a solution of toluoyl derivative (1 eq) in CH₃CN (1.5 mL/mmol oftoluoyl derivative) was added N-bromosuccinimide (1.2 eq) and2,2′-azobis(2-methylpropionitrile) (0.02 eq). The reaction mixture washeated at 70° C. for 1 to 48 hours. The reaction mixture was then cooledto RT and water was added and the mixture was concentrated under vacuum.The residue was taken up in DCM and washed with a 10% aqueous solutionof NaHCO₃ and brine, dried over MgSO₄ and concentrated under vacuum.Purification by flash chromatography or recrystallization afforded theexpected benzyl bromide derivative.

Procedure 14: Amide Formation

To a solution of the carboxylic acid derivative (1 eq) in DCM (40mL/mmol) was added oxalyl chloride (3 eq) and DMF (0.04 eq) in DCM (10mL) and the resulting mixture was stirred at RT for 1 hour. The solutionwas evaporated to dryness, the residue taken up in THF (10 mL/mmol) andthen added to a mixture of the amine derivative (1 eq) and DIEA (4 eq)in THF (8 mL/mmol). The reaction mixture was stirred at RT for 2 to 24hours, filtered through a SPE NH₂ column and rinsed with ACN. Afterevaporation of the solvents, the crude product was purified by flashchromatography affording the title compound.

Procedure 15: Suzuki Cross-Coupling Reaction

A mixture of the aryl bromide (1 eq), the boronic acid or esterderivative (1.2 to 2.0 eq), cesium fluoride (3 eq), palladium acetate(0.05 to 0.2 eq) and 2-dicyclohexylphosphino-2′,6′-dimethoxybiphenyl(0.1 to 0.4 eq) was prepared in dioxane (10 mL/g of aryl bromide) andwater (5 mL/g of aryl bromide) under nitrogen atmosphere. The resultingmixture was heated at a temperature comprised between 80° C. to refluxfor 1 to 15 hours. The reaction mixture was diluted with MTBE (40 mL/gof aryl bromide) and the aqueous layer was removed. The organic layerwas dried (MgSO₄ or Na₂SO₄) and concentrated under vacuum. The residuewas purified by flash chromatography.

Intermediate 5: 2′-methyl-2-(trifluoromethyl)biphenyl-4-carboxylic acid

Step 1: Methyl 4-bromo-3-(trifluoromethyl)benzoate

To a suspension of 4-bromo-3-(trifluoromethyl)benzoic acid (Acceledev000625, 15.0 g, 55.8 mmol) in MeOH (300 mL) at RT was added dropwisethionyl chloride (16.2 mL, 223.0 mmol) over 15 minutes. The reactionmixture was stirred at RT for 12 hours. The reaction mixture wasconcentrated and the crude residue was diluted with EtOAc (500 mL). Theorganic layer was washed with a saturated aqueous solution of NaHCO₃(200 mL), water (200 mL), brine (200 mL), dried over MgSO₄ andconcentrated affording the title compound as an orange solid (14.8 g,94%). ¹H NMR (DMSO-d₆, 300 MHz) δ 8.26 (m, 1H), 8.14 (m, 2H), 3.93 (s,3H). HPLC (Method A) Rt 4.71 min (Purity: 99.0%).

Step 2: Methyl 2′-methyl-2-(trifluoromethyl)biphenyl-4-carboxylate

Methyl 4-bromo-3-(trifluoromethyl)benzoate (6.0 g, 21.2 mmol),o-tolylboronic acid (3.17 g, 23.3 mmol), potassium carbonate (14.7 g,106.0 mmol), tetrakis(triphenylphosphine)palladium(0) (2.45 g, 2.12mmol) were taken up in toluene (30 mL) and water (30 mL) under N₂atmosphere. The reaction mixture was refluxed for 3 hours. The reactionmixture was cooled to RT, filtered over a pad of celite and rinsed withtoluene (200 mL). The filtrate was concentrated to afford a brown oilwhich was taken up in EtOAc (200 mL). The organic layer was washed witha saturated aqueous solution of NaHCO₃ (50 mL), water (50 mL) and brine(50 mL), dried over MgSO₄ and concentrated affording the title compoundas a brown oil (6.4 g, quantitative). HPLC (Method A) Rt 5.33 min(Purity: 60%).

Step 3: 2′-methyl-2-(trifluoromethyl)biphenyl-4-carboxylic acid

A solution of methyl 2′-methyl-2-(trifluoromethyl)biphenyl-4-carboxylate(5.0 g, 17.0 mmol) in EtOH (150 mL) at RT was treated with a 5N aqueoussolution of sodium hydroxide (10.2 mL, 51 mmol). The reaction mixturewas stirred at 60° C. for 2 hours. The reaction mixture was concentratedto give a brown solid which was taken up in water (300 mL) and theaqueous phase was washed twice with EtOAc. The aqueous phase wasacidified with a concentrated aqueous solution of HCl (pH-2), then itwas concentrated until precipitation (half of the volume). Thesuspension was filtered affording the title compound as a beige solid.¹H NMR (DMSO-d₆, 300 MHz) δ 13.55 (br s, 1H), 8.31 (s, 1H), 8.25 (d,J=7.9 Hz, 1H), 7.50 (d, J=7.9 Hz, 1H), 7.37-7.12 (m, 4H), 1.99 (s, 3H).LC/MS (Method A): 278.9 (M−H)⁻. HPLC (Method A) Rt 4.57 min (Purity:98.7%).

Intermediate 16: tert-butyl3-[{3-[amino(hydroxyimino)methyl]benzyl}(methyl)amino]propanoate

Step 1: 3-[(Methylamino)methyl]benzonitrile

To a stirred solution of methylamine (40% in water, 200 mL) under N₂,was added 3-(bromomethyl)benzonitrile (10 g, 51 mmol) slowly in portionsover 10 minutes at 0° C. After being stirred at RT for 3 hours, thereaction mixture was extracted with DCM. Then the organic layer waswashed with brine and dried over sodium sulphate and concentrated underreduced pressure. The residue was purified by chromatography (silica,pet ether/EtOAc) to afford the title compound as a pale yellow liquid(6.1 g, 82%). ¹H NMR (DMSO-d₆, 400 MHz) δ 7.74 (s, 1H), 7.63-7.68 (m,2H), 7.48-7.52 (m, 1H), 3.66 (s, 2H), 2.22 (s, 3H).

Step 2: tert-butyl 3-[(3-cyanobenzyl)(methyl)amino]propanoate

To a stirred mixture of 3-[(methylamino)methyl]benzonitrile (6.1 g, 41.7mmol) and sodium bicarbonate (7.0 g, 83.4 mmol) in ACN (70 mL) was addeddropwise tert-butyl 3-bromo-propanoate (7 mL, 41.7 mol). After beingstirred at RT for 12 hours, the reaction mixture was filtered and thefiltrate was concentrated under reduced pressure. The residue waspurified by chromatography (silica, pet ether/EtOAc) to afford the titlecompound as a pale yellow liquid. ¹H NMR (DMSO-d₆, 400 MHz) δ 7.70 (s,2H), 7.60 (m, 1H), 7.51 (m, 1H), 3.49 (s, 2H), 2.58 (m, 2H), 2.36 (m,2H), 2.09 (s, 3H), 1.38 (s, 9H).

Step 3: tert-butyl3-[{3[amino(hydroxyimino)methyl]benzyl}(methyl)amino]propanoate

The title compound was prepared following the general procedure 1,starting from tert-butyl 3-[(3-cyanobenzyl)(methyl)amino]propanoate. Itwas obtained as a white gummy solid (4.50 g, 84%). ¹H NMR (DMSO-d₆, 400MHz) δ 9.56 (s, 1H), 7.52 (m, 2H), 7.29 (m, 2H), 5.74 (s, 2H), 3.44 (s,2H), 2.57 (m, 2H), 2.36 (m, 2H), 2.09 (s, 3H), 1.37 (s, 9H). LC/MS(Method B): 308.2 (M+H)⁺. HPLC (Method B) Rt 5.18 min (Purity: 96.5%).

Intermediate 21:tert-butyl[{3-[amino(hydroxyimino)methyl]benzyl}(methyl)amino]acetate

Step 1: tert-butyl[(3-cyanobenzyl)(methyl)amino]acetate

To a stirred solution of sarcosine tert-butyl ester hydrochloride (8.1g, 44.9 mmol) and triethylamine (17 mL, 122.4 mmol) in ACN (100 mL) wasadded 3-(bromomethyl)benzonitrile (8.0 g, 40.8 mmol) portionwise over aperiod of 10 minutes at 0° C. After being stirred at RT for 3 hours, thereaction mixture was poured into water and extracted with DCM. Then theorganic layer was washed with brine, dried over sodium sulphate andconcentrated under reduced pressure to afford the title compound as apale green liquid (9.0 g, 85%). ¹H NMR (DMSO-d₆, 400 MHz) δ 7.72 (2H,m), 7.64 (1H, m), 7.54 (1H, m), 3.66 (2H, s), 3.18 (2H, s), 2.22 (3H,s), 1.41 (9H, s).

Step 2:tert-butyl[{3-[amino(hydroxyimino)methyl]benzyl}(methyl)amino]acetate

The title compound was prepared according the general procedure 1,starting from tert-butyl[(3-cyanobenzyl)(methyl)amino]acetate. It wasobtained as a white powder (8.5 g, 84%). ¹H NMR (DMSO-d₆, 400 MHz) δ9.57 (1H, s), 7.59 (1H, s), 7.53 (1H, m), 7.30 (2H, m), 5.75 (2H, s),3.61 (2H, s), 3.15 (2H, s), 2.23 (3H, s), 1.41 (9H, s). LC/MS (MethodB): 294.0 (M+H)⁺. HPLC (Method A) Rt 3.31 min (Purity: 97.5%).

Intermediate 22: tert-butylN-{3-[amino(hydroxyimino)methyl]benzyl}-N-(tert-butoxycarbonyl)-beta-alaninate

Step 1: tert-butyl 3-[(3-cyanobenzyl)amino]propanoate

To a stirred solution of β-alanine tert-butyl ester hydrochloride (7.1g, 39.2 mmol) and triethylamine (14.2 mL, 107 mmol) in dry acetonitrile(100 mL) was added 3-(bromomethyl)benzonitrile (7.0 g, 35.7 mmol) inportions over 10 minutes at 0° C. After being stirred at RT for 5 hours,the reaction mixture was diluted with water (100 mL) and extracted withdichloromethane (2×100 mL) The organic layers were combined, dried usingsodium sulphate and concentrated under reduced pressure. The residue waspurified by flash chromatography affording the title compound as a paleyellow liquid. ¹H NMR (DMSO-d₆, 400 MHz) δ 7.75 (s, 1H), 7.66 (m, 2H),7.50 (m, 1H), 3.71 (s, 2H), 2.64 (t, 2H), 2.32 (m, 2H), 1.37 (s, 9H).

Step 2: tert-butylN-(tert-butoxycarbonyl)-N-(3-cyanobenzyl)-beta-alaninate

To a solution of tert-butyl 3-[(3-cyanobenzyl)amino]propanoate (3.7 g,14.2 mmol) in anhydrous THF (100 mL) was added di-tert-butyl dicarbonate(3.1 g, 14.2 mmol). The resulting mixture was heated at 50° C. for 6hours. The reaction mixture was then concentrated under reduced pressureaffording the title compound as a pale yellow liquid (5.0 g, 98%). ¹HNMR (DMSO-d₆, 400 MHz) δ 7.73 (t, 1H), 7.63 (s, 1H), 7.55 (d, 2H), 4.41(s, 2H), 3.31 (t, 2H), 2.40 (t, 2H), 1.36 (m, 18H).

Step 3: tert-butylN-{3-[amino(hydroxyimino)methyl]benzyl}-N-(tert-butoxycarbonyl)-beta-alaninate

The title compound was prepared following the general procedure 1,starting from tert-butylN-(tert-butoxycarbonyl)-N-(3-cyanobenzyl)-beta-alaninate (5.0 g, 13.8mmol). It was isolated as a pale yellow oil (5.1 g, 93%). ¹H NMR(DMSO-d₆, 400 MHz) δ 9.61 (s, 1H), 7.54 (m, 2H), 7.33 (m, 1H), 7.19 (d,1H), 5.78 (s, 2H), 4.38 (s, 2H), 3.33 (m, 2H), 2.40 (m, 2H), 1.33-1.42(m, 18H). LC/MS (Method B): 394.0 (M+H)⁺. HPLC (Method A) Rt 3.47 min(Purity: 97.7%). HPLC (Method E) Rt 3.71 min (Purity: 97.2%).

Intermediate 23: tert-butylN-{5-[amino(hydroxyimino)methyl]-2-fluorobenzyl}-N-methyl-beta-alaninate

Step 1: tert-butyl N-(5-cyano-2-fluorobenzyl)-N-methyl-beta-alaninate

The title compound was prepared following the general procedure 10starting from 5-cyano-2-fluorobenzylbromide (1.5 g, 7.0 mmol) andtert-butyl N-methyl-beta-alaninate (1.3 g, 8.4 mmol, prepared asdescribed in Biorg. Med. Chem. (11) 2003, 3083-3099). It was isolated asa pale yellow oil (1.9 g, 92%). ¹H NMR (CDCl₃, 300 MHz) δ 7.79 (dd,J=6.7, 2.1 Hz, 1H), 7.57-7.52 (m, 1H), 7.11 (m, 1H), 3.57 (s, 2H), 2.74(t, J=7.1 Hz, 2H), 2.43 (t, J=7.1 Hz, 2H), 2.22 (s, 3H), 1.45 (s, 9H).LC/MS (Method B): 293.2 (M+H)⁺. HPLC (Method A) Rt 2.44 min (Purity:86.4%).

Step 2: tert-butylN-{5-[amino(hydroxyimino)methyl]-2-fluorobenzyl}-N-methyl-beta-alaninate

The title compound was prepared following the general procedure 1starting from tert-butylN-(5-cyano-2-fluorobenzyl)-N-methyl-beta-alaninate (1.39 g, 4.75 mmol).It was isolated as a colorless oil (1.6 g, quantitative). ¹H NMR (CDCl₃,300 MHz) δ 7.73 (dd, J=7.0, 2.3 Hz, 1H), 7.58-7.53 (m, 1H), 7.03 (dd,J=9.5, 8.7 Hz, 1H), 5.11 (br s, 2H), 3.59 (s, 2H), 2.69 (t, J=7.0 Hz,2H), 2.45 (t, J=7.0 Hz, 2H), 2.28 (s, 3H), 1.45 (s, 9H). LC/MS (MethodB): 324.3 (M−H)⁻, 326.2 (M+H)⁺.

Intermediate 24: tert-butylN-{3-[amino(hydroxyimino)methyl]-5-fluorobenzyl}-N-methylglycinate

Step 1: 3-(bromomethyl)-5-fluorobenzonitrile

A solution of 3-fluoro-5-methylbenzonitrile (Hognda Trading Co) (100 g,0.74 mol) was prepared in ACN (1 L) under nitrogen atmosphere.N-Bromosuccinimide (105 g, 0.59 mol) and AIBN (2.4 g, 0.014 mol) wereadded and the reaction mixture was heated at 70° C. for 90 minutes. Thereaction mixture was concentrated under vacuum. The residue was dilutedin DCM, cooled at 0° C. and stirred for 15 minutes. The precipitatedsuccinimide was filtered off and the filtrate was concentrated to give ayellow oil. The crude product was taken up with pet ether (200 mL),cooled at −20° C. and stirred for 30 minutes. The precipitate wasfiltered off and dried under vacuum to give the title compound as awhite solid. ¹H NMR (DMSO-d₆, 400 MHz) δ 7.83 (m, 2H) 7.73 (m, 1H), 4.72(s, 2H). HPLC (Method A) Rt 4.17 min (Purity: 99.4%).

Step 2: tert-butyl N-(3-cyano-5-fluorobenzyl)-N-methylglycinate

The title compound was prepared according the general procedure 10,starting from 3-(bromomethyl)-5-fluorobenzonitrile and sarcosinetert-butyl ester hydrochloride. It was isolated as a yellow oil (3.5 g,98%). ¹H NMR (DMSO-d₆, 400 MHz) δ 7.74 (m, 1H), 7.63 (m, 1H), 7.54 (m,1H), 3.70 (brs, 2H), 3.22 (brs, 2H), 2.24 (s, 3H), 1.42 (s, 9H). LC/MS(Method B) 262.2 (M+H)⁺. HPLC (Method A) Rt 2.42 min (Purity: 93.3%).

Step 3: tert-butylN-{3-[amino(hydroxyimino)methyl]-5-fluorobenzyl}-N-methylglycinate

The title compound was prepared according general procedure 2, startingfrom tert-butyl N-(3-cyano-5-fluorobenzyl)-N-methylglycinate. It wasisolated as a yellow solid. ¹H NMR (DMSO-d₆, 400 MHz) δ 9.75 (br s, 1H),7.49 (s, 1H), 7.34 (m, 1H), 7.13 (m, 1H), 5.87 (brs, 2H), 3.65 (s, 2H),3.19 (s, 2H), 2.25 (s, 3H), 1.43 (s, 9H). LC/MS (Method B): 311.9(M+H)⁺, 310.0 (M−H⁻). HPLC (Method A) Rt 1.89 min (Purity: 53%).

Intermediate 25: tert-butylN-{3-[amino(hydroxyimino)methyl]-5-fluorobenzyl}-N-methyl-beta-alaninate

Step 1: tert-butyl N-(3-cyano-5-fluorobenzyl)-methyl-beta-alaninate

The title compound was prepared according the general procedure 10,starting from 3-(bromomethyl)-5-fluorobenzonitrile (Intermediate 24 Step1, 750 mg, 3.5 mmol) and tert-butyl N-methyl-beta-alaninate (670 mg, 4.2mmol, prepared as described in Biorg. Med. Chem. (11) 2003, 3083-3099).It was isolated as a pale yellow oil. ¹H NMR (DMSO-d₆, 300 MHz) δ 7.73(ddd, J=8.6, 2.6, 1.4 Hz, 1H), 7.61 (s, 1H), 7.56-7.45 (m, 1H), 3.53 (s,2H), 2.60 (t, J=6.7 Hz, 2H), 2.38 (t, J=6.7 Hz, 2H), 2.12 (s, 3H), 1.40(s, 9H). LC/MS (Method B): 293.3 (M+H)⁺. HPLC (Method A) Rt 2.61 min(Purity: 96.2%).

Step 2: tert-butylN-{3-[amino(hydroxyimino)methyl]-5-fluorobenzyl}-N-methyl-beta-alaninate

The title compound was prepared following the general procedure 2,starting from tert-butylN-(3-cyano-5-fluorobenzyl)-N-methyl-beta-alaninate (495 mg, 1.7 mmol).It was isolated as a yellow oil (524 mg, 95%). ¹H NMR (DMSO-d₆, 300 MHz)δ 9.73 (s, 1H), 7.45 (s, 1H), 7.38-7.28 (m, 1H), 7.11 (d, J=9.7 Hz, 1H),5.85 (s, 2H), 3.48 (s, 2H), 2.60 (t, J=6.9 Hz, 2H), 2.38 (t, J=6.9 Hz,2H), 2.12 (s, 3H), 1.39 (s, 9H). LC/MS (Method B): 324.3 (M−H)⁻.

Intermediate 29:3-{[(2,3-dihydroxypropyl)(methyl)amino]methyl}-N′-hydroxybenzenecarboximidamide

Step 1: 3-{[(2,3-dihydroxypropyl)(methyl)amino]methyl}benzonitrile

The title compound was prepared following the general procedure 10,starting from 3-(bromomethyl)benzonitrile and3-(methylamino)-1,2-propanediol. It was isolated as a colorless oil (920mg, 82%). LC/MS (Method B): 221.1 (M+H)⁺.

Step 2:3-{[(2,3-dihydroxypropyl)(methyl)amino]methyl}-N′-hydroxybenzenecarboximidamide

The title compound was prepared following the general procedure 1,starting from3-{[(2,3-dihydroxypropyl)(methyl)amino]methyl}benzonitrile. It wasisolated as a colorless oil. LC/MS (Method B): 254.1 (M+H)⁺.

Intermediate 30:3-{[bis(2-hydroxyethyl)amino]methyl}-N′-hydroxybenzenecarboximidamide

Step 1: 3-{[bis(2-hydroxyethyl)amino]methyl}benzonitrile

The title compound was prepared following the general procedure 10,starting from 3-(bromomethyl)benzonitrile and diethanolamine. It wasobtained as a colorless oil (820 mg, 73%). LC/MS (Method B): 219.1(M−H)⁻, 221.1 (M+H)⁺.

Step 2:3-{[bis(2-hydroxyethyl)amino]methyl}-N′-hydroxybenzenecarboximidamide

The title compound was prepared following the general procedure 2,starting from 3-{[bis(2-hydroxyethyl)amino]methyl}benzonitrile. It wasobtained as a colorless oil.

Intermediate 31: tert-butylN-{4-[amino(hydroxyimino)methyl]benzyl}-N-methylglycinate

Step 1: tert-butyl N-(4-cyanobenzyl)-N-methylglycinate

A mixture of alpha-bromo-p-tolunitrile (10.0 g, 51 mmol), sarcosinetert-butyl ester hydrochloride (10.2 g, 56 mmol) and potassium carbonate(21.2 g, 153 mmol) in acetone (100 mL) was stirred at 60° C. overnight.Resulting suspension was filtered off and salts were washed with acetone(2×150 mL). The filtrate was concentrated under reduced pressure. Theresidue was diluted with water (200 mL) and extracted with EtOAc (2×150mL). The combined organic layers were washed with water (200 mL), dried(MgSO₄) and the solvents were removed under reduced pressure to give thetitle compound as a yellow oil used without further purification (11.7g, 88%). ¹H NMR (DMSO-d₆, 300 MHz) δ 7.79 (d, J=8.4 Hz, 2H), 7.51 (d,J=8.4 Hz, 2H), 3.71 (s, 2H), 3.20 (s, 2H), 2.23 (s, 3H), 1.42 (s, 9H).LC/MS (Method B): 260.9 (M+H)⁺.

Step 2: tert-butylN-{4-[amino(hydroxyimino)methyl]benzyl}-N-methylglycinate

A 50% aqueous solution of hydroxylamine (13.25 mL, 225 mmol) was addedat once into a solution of tert-butylN-(4-cyanobenzyl)-N-methylglycinate (11.70 g, 45 mmol) in EtOH (75 mL).The reaction mixture was stirred at RT overnight. The solvents wereremoved under reduced pressure to give a white solid. The solid wassuspended in water (200 mL) and stirred for 20 minutes, then filteredoff, washed with water and dried to give the title compound as a whitepowder (12.54 g, 95%). ¹H NMR (DMSO-d₆, 300 MHz) δ 9.58 (s, 1H), 7.62(d, J=8.3 Hz, 2H), 7.28 (d, J=8.3 Hz, 2H), 5.78 (s, 2H), 3.62 (s, 2H),3.15 (s, 2H), 2.24 (s, 3H), 1.42 (s, 9H). LC/MS (Method B): 294.0(M+H)⁺.

Intermediate 32: tert-butylN-{4-[amino(hydroxyimino)methyl]-2-fluorobenzyl}-N-methylglycinate

Step 1: tert-butyl N-(4-cyano-2-fluorobenzyl)-N-methylglycinate

A mixture of 4-cyano-2-fluorobenzyl bromide (250 g, 1.17 mol), sarcosinetert-butyl ester hydrochloride (212.2 g, 1.17 mol) and potassiumcarbonate (484 g, 3.50 mol) in acetone (2 L) was stirred 24 hours at 50°C. and 18 hours at RT. The reaction mixture was filtered and thefiltrate was concentrated under vacuum. The residue was taken up withwater (1 L) and extracted with ethyl acetate (2×1 L). The combinedorganic layers were washed with water (1 L), dried (MgSO₄) andconcentrated under vacuum to give the title compound as a yellow-brownoil (325 g, quantitative). ¹H NMR (DMSO-d₆, 300 MHz) δ 7.82 (dd, J=9.9,1.4 Hz, 1H), 7.70 (dd, J=7.9, 1.5 Hz, 1H), 7.64 (m, 1H), 3.78 (s, 2H),3.23 (s, 2H), 2.27 (s, 3H), 1.42 (s, 9H). LC/MS (Method B): 279.2(M+H)⁺.

Step 2: tert-butylN-{4-[amino(hydroxyimino)methyl]-2-fluorobenzyl}-N-methylglycinate

A 50% aqueous solution of hydroxylamine (345 mL, 5.85 mol) was addeddropwise over 10 minutes into a solution of tert-butylN-(4-cyano-2-fluorobenzyl)-N-methylglycinate (325 g, 1.17 mol) in EtOH(2.1 L). The reaction mixture was stirred at RT for 20 hours. Solventswere concentrated under vacuum and water (2 L) was added. The resultingsuspension was filtered off, washed with water (1 L) and dried undervacuum to give title compound as white powder (323 g, 89%). ¹H NMR(DMSO-d₆, 300 MHz) δ 9.74 (s, 1H), 7.51 (dd, J=8.0, 1.6 Hz, 1H),7.45-7.36 (m, 2H), 5.87 (s, 2H), 3.70 (s, 2H), 3.17 (s, 2H), 2.26 (s,3H), 1.42 (s, 9H). LC/MS (Method B): 312.3 (M+H)⁺.

Intermediate 33: tert-butylN-{4-[amino(hydroxyimino)methyl]-2-bromobenzyl}-N-methylglycinate

Step 1: tert-butyl N-(2-bromo-4-cyanobenzyl)-N-methylglycinate

The title compound was prepared following the general procedure 10,starting from 2-bromo-4-cyanobenzyl bromide (Carbocor) and sarcosinetert-butyl ester hydrochloride. It was obtained as a white solid. ¹H NMR(DMSO-d₆, 300 MHz) δ 8.17 (d, J=1.6 Hz, 1H), 7.88 (dd, J=8.0, 1.6 Hz,1H), 7.69 (d, J=8.0 Hz, 1H), 3.82 (s, 2H), 3.31 (s, 2H), 2.31 (s, 3H),1.43 (s, 9H). LC/MS (Method B): 338.9, 340.9 (M+H)⁺. HPLC (Method A) Rt3.10 min (Purity: 99.6%).

Step 2: tert-butylN-{4-[amino(hydroxyimino)methyl]-2-bromobenzyl}-N-methylglycinate

The title compound was prepared following the general procedure 2,starting from tert-butyl N-(2-bromo-4-cyanobenzyl)-N-methylglycinate. Itwas isolated as a colorless oil (303 mg, 97%). ¹H NMR (DMSO-d₆, 300 MHz)δ 9.74 (s, 1H), 7.87 (d, J=1.7 Hz, 1H), 7.67 (dd, J=8.0, 1.7 Hz, 1H),7.47 (d, J=8.0 Hz, 1H), 5.88 (s, 2H), 3.75 (s, 2H), 3.26 (s, 2H), 2.31(s, 3H), 1.43 (s, 9H). LC/MS (Method B): 371.9, 373.9 (M+H)⁺. HPLC(Method A) Rt 1.53 min (Purity: 98.6%).

Intermediate 34: methyl 4-[amino(hydroxyimino)methyl]-2-chlorobenzoate

Step 1: methyl 4-bromo-2-chlorobenzoate

Thionyl chloride (1.23 mL, 17.0 mmol) was added dropwise into a solutionof 4-bromo-2-chlorobenzoic acid (Combi-Blocks CA-4187, 1.0 g, 4.25 mmol)in MeOH (20 mL) at 0° C. The reaction mixture was then stirred at RTuntil completion. The reaction mixture was concentrated under reducedpressure. The residue was dissolved in EtOAc and washed with a saturatedaqueous solution of NaHCO₃ and brine. The organic layer was dried(MgSO₄) and concentrated under vacuum to give the title compound (871mg, 82%). ¹H NMR (CDCl₃, 300 MHz) δ 7.72 (d, J=8.4 Hz, 1H), 7.64 (d,J=1.9 Hz, 1H), 7.46 (dd, J=8.4, 1.9 Hz, 1H), 3.93 (s, 3H). HPLC (MethodA) Rt 4.22 min (Purity: 98.5%).

Step 2: methyl 2-chloro-4-cyanobenzoate

A mixture of triphenylphosphine polymer bound (239 mg, 0.72 mmol) andpalladium(II) acetate (75 mg, 0.33 mmol) was prepared in DMF (12 mL)under nitrogen atmosphere and stirred at RT for 2 hours. Zinc cyanide(561 mg, 4.77 mmol) and methyl 4-bromo-2-chlorobenzoate (1.19 g, 4.77mmol) were added and the resulting mixture was heated under microwaveirradiation at 140° C. for 50 minutes. The reaction mixture was filteredand the resin was washed with Et₂O (3×10 mL). The combined organiclayers were washed with water (3×5 mL) and brine (10 mL), dried (MgSO₄)and concentrated under vacuum to give the title compound as a whitepowder. ¹H NMR (DMSO-d₆, 300 MHz) δ 8.23 (m, 1H), 7.96 (m, 2H), 3.90 (s,3H). HPLC (Method A) Rt 3.39 min (Purity: 97.6%).

Step 3: methyl 4-[amino(hydroxyimino)methyl]-2-chlorobenzoate

The title compound was prepared according the general procedure 1,starting from methyl 2-chloro-4-cyanobenzoate. It was obtained as abeige solid (631 mg, quantitative). ¹H NMR (DMSO-d₆, 300 MHz) δ 10.02(br s, 1H), 7.84 (d, J=1.6 Hz, 1H), 7.83 (d, J=8.2 Hz, 1H), 7.75 (dd,J=8.2, 1.6 Hz, 1H), 6.03 (br s, 2H), 3.86 (s, 3H). LC/MS (Method B):227.2 (M−H)⁻, 229.1 (M+H)⁺. HPLC (Method A) Rt 1.44 min (Purity: 63.4%).

Intermediate 35: methyl 4-[amino(hydroxyimino)methyl]-2-fluorobenzoate

Step 1: methyl 4-cyano-2-fluorobenzoate

Oxalyl chloride (9.0 mL, 106.6 mmol) and DMF (0.5 mL) were added into asuspension of 4-cyano-2-fluorobenzoic acid (ABCR, 16 g, 96.9 mmol) inanhydrous DCM (300 mL) The resulting mixture was stirred at RT, and thenevaporated under reduced pressure. The residue was taken up withanhydrous THF (150 mL) and added dropwise into a solution of methanol(50 mL) and triethylamine (25.8 mL, 193.8 mmol) at 4° C. The reactionmixture was diluted with a 0.1N aqueous solution of HCl (200 mL) andextracted with EtOAc (3×100 mL). The combined organic layers were washedwith a semi-saturated aqueous solution of NaHCO₃ (200 mL) and water (200mL), dried (MgSO₄) and concentrated under vacuum to give the titlecompound as a pale yellow solid (17.8 g, quantitative). HPLC (Method A)Rt 2.81 min (Purity: 93.9%).

Step 2: methyl 4-[amino(hydroxyimino)methyl]-2-fluorobenzoate

The title compound was prepared according the general procedure 1,starting from methyl 4-cyano-2-fluorobenzoate. It was obtained as whitesolid. ¹H NMR (DMSO-d₆, 300 MHz) δ 10.09 (s, 1H), 7.92 (m, 1H),7.70-7.62 (m, 2H), 6.05 (s, 2H), 3.89 (s, 3H). LC/MS (Method B): 210.9(M−H)⁻, 212.9 (M+H)⁺. HPLC (Method A), Rt 0.97 min (purity: 100%).

Intermediate 36: tert-butylN-{4-[amino(hydroxyimino)methyl]benzyl}-N-(tert-butoxycarbonyl)glycinate

Step 1: tert-butyl N-(4-cyanobenzyl)glycinate

The title compound was prepared following the general procedure 10,starting from 4-cyanobenzyl bromide and tert-butyl glycinate. It wasisolated as a colorless oil. ¹H NMR (DMSO-d₆, 300 MHz) δ 7.78 (d, J=8.3Hz, 2H), 7.52 (d, J=8.3 Hz, 2H), 3.79 (s, 2H), 3.19 (s, 2H), 2.60 (br s,1H), 1.41 (s, 9H). HPLC (Method A) Rt 2.17 min (Purity: 97.2%).

Step 2: tert-butyl N-(tert-butoxycarbonyl)-N-(4-cyanobenzyl)glycinate

A mixture of tert-butyl N-(4-cyanobenzyl)glycinate (1.1 g, 4.5 mmol),di-tert-butyl dicarbonate (1.1 g, 4.9 mmol) and N-ethyldiisopropylamine(1.1 mL, 6.7 mmol) was prepared in DCM (22 mL) and stirred at RT for 3.5hours. The reaction mixture was diluted with DCM and washed with asaturated aqueous solution of NaHCO₃ (2×) and brine. The organic layerwas dried (MgSO₄) and concentrated to afford the title compound as acolorless oil (1.2 g, 77%). ¹H NMR (DMSO-d₆, 300 MHz) δ 7.84-7.78 (m,2H), 7.48 (d, J=8.0 Hz, 2H), 4.50-4.45 (m, 2H), 3.93-3.83 (m, 2H),1.41-1.29 (m, 18H). LC/MS (Method B): 347.1 (M+H)⁺. HPLC (Method A) Rt5.10 min (Purity: 100.0%).

Step 3: tert-butylN-{4-[amino(hydroxyimino)methyl]benzyl}-N-(tert-butoxycarbonyl)glycinate

The title compound was prepared following the general procedure 2,starting from tert-butylN-(tert-butoxycarbonyl)-N-(4-cyanobenzyl)glycinate. It was obtained as acolorless oil (1.26 g, 97%). ¹H NMR (DMSO-d₆, 300 MHz) δ 9.59 (s, 1H),7.65-7.60 (m, 2H), 7.26 (d, J=8.3 Hz, 2H), 5.77 (s, 2H), 4.40 (s, 2H),3.84-3.75 (m, 2H), 1.41-1.34 (m, 18H). LC/MS (Method B): 380.1 (M+H)⁺.HPLC (Method A) Rt 3.31 min (Purity: 98.1%).

Intermediate 37: tert-butylN-{3-[amino(hydroxyimino)methyl]benzyl}-N-ethyl-beta-alaninate

Step 1: 3-[(ethylamino)methyl]benzonitrile

3-(Bromomethyl)benzonitrile (20.0 g, 0.1 mol) was added portion wiseinto a 2M solution of ethylamine in THF (150 mL) at 0° C. After 5 hours,the reaction mixture was concentrated under vacuum. The residue wastaken up with water and extracted with DCM. The organic layer was driedover sodium sulphate and evaporated under reduced pressure to afford thetitle compound as yellow liquid (12 g, 73%). ¹H NMR (DMSO-d₆, 400 MHz) δ7.75 (s, 1H), 7.67 (d, 2H), 7.50 (m, 1H), 3.71 (s, 2H), 2.46 (m, 2H),0.99 (m, 3H).

Step 2: tert-butyl N-(3-cyanobenzyl)-N-ethyl-beta-alaninate

A mixture of 3-[(ethylamino)methyl]benzonitrile (5 g, 31 mmol) andtert-butyl acrylate (4.6 mL, 31 mmol) was heated neat at 80° C. for 12hours. The crude mixture was purified by flash chromatography (silica,pet ether/EtOAc) to afford the title compound as colorless oil. ¹H NMR(DMSO-d₆, 400 MHz) δ 7.69 (m, 2H), 7.62 (d, J=7.8 Hz, 1H), 7.50 (m, 1H),3.56 (s, 2H), 2.64 (t, 2H), 2.40 (m, 2H), 2.32 (t, 2H), 1.39 (s, 9H),0.93 (t, 3H).

Step 3: tert-butylN-{3-[amino(hydroxyimino)methyl]benzyl}-N-ethyl-beta-alaninate

The title compound was prepared according the general procedure 2,starting from tert-butyl N-(3-cyanobenzyl)-N-ethyl-beta-alaninate. Itwas isolated as a colorless oil (5.2 g, 86%). ¹H NMR (DMSO-d₆, 400 MHz)δ 9.59 (1H, s), 7.59 (2H, m), 7.52 (d, J=5.8 Hz, 1H), 7.29 (d, J=5.8 Hz,1H), 5.74 (s, 2H), 3.51 (s, 2H), 2.66 (t, 2H), 2.41 (m, 2H), 2.32 (t,2H), 1.38 (s, 9H), 0.95 (t, 3H). LC/MS (Method A) 322.0 (M+H)⁺. HPLC(Method A) Rt 2.09 min (Purity: 97.2%).

Intermediate 38: tert-butylN-{3-[amino(hydroxyimino)methyl]benzyl}-N-(2-hydroxyethyl)-beta-alaninate

Step 1: 3-{[(2-hydroxyethyl)amino]methyl}benzonitrile

3-(Bromomethyl)benzonitrile (20.0 g, 0.125 mol) was added portion wiseinto a mixture of ethanolamine (11.4 g, 0.19 mol) and sodium bicarbonate(21.0 g, 0.25 mol) in ACN (200 mL) at 0° C. The resulting mixture wasstirred for 4 hours. The reaction mixture was concentrated under vacuum.The residue was taken up with water and extracted with DCM (200 mL). Theorganic layer was dried over sodium sulfate and evaporated under reducedpressure. The residue was purified by chromatography to afford the titlecompound as white solid (9.8 g, 54%). ¹H NMR (DMSO-d₆, 400 MHz) δ 7.76(s, 1H), 7.67 (m, 2H), 7.51 (m, 1H), 4.47 (brs, 1H), 3.74 (s, 2H), 3.44(m, 2H), 2.51 (m, 2H), 2.12 (brs, 1H).

Step 2: tert-butyl N-(3-cyanobenzyl)-N-(2-hydroxyethyl)-beta-alaninate

Tert-butyl acrylate (3.56 ml, 29 mmol) was added into a solution of3-{[(2-hydroxyethyl)amino]methyl}benzonitrile (4.9 g, 28 mmol) inanhydrous DMF (5 mL) under nitrogen atmosphere. The resulting mixturewas heated at 80° C. for 12 hours. The reaction mixture was diluted withwater (100 mL) and extracted with ethyl acetate (100 mL). The organiclayer was dried over sodium sulfate and concentrated under reducedpressure to afford the title compound as colorless liquid. ¹H NMR(DMSO-d₆, 400 MHz) δ 7.75 (s, 1H), 7.69 (d, J=7.6 Hz, 1H), 7.62 (d,J=7.9 Hz, 1H), 7.50 (m, 1H), 4.36 (t, 1H), 3.62 (s, 2H), 3.43 (m, 2H),2.69 (t, 2H), 2.47 (m, 2H), 2.34 (t, 2H), 1.38 (s, 9H).

Step 3: tert-butylN-{3-[amino(hydroxyimino)methyl]benzyl}-N-(2-hydroxyethyl)-beta-alaninate

The title compound was prepared following the general procedure 2,starting from tert-butylN-(3-cyanobenzyl)-N-(2-hydroxyethyl)-beta-alaninate. It was isolated asa colorless oil (4.0 g, 87%). ¹H NMR (DMSO-d₆, 400 MHz) δ 9.57 (s, 1H),7.58 (s, 1H), 7.51 (t, 1H), 7.28 (d, 2H), 5.74 (s, 2H), 4.30 (t, 1H),3.57 (s, 2H), 3.41 (m, 2H), 2.70 (t, 2H), 2.47 (m, 2H), 2.34 (t, 2H),1.37 (s, 9H). LC/MS (Method A) 338.1 (M+H)⁺. HPLC (Method A) Rt 1.89 min(Purity: 98.2%).

Intermediate 39: tert-butylN-{4-[amino(hydroxyimino)methyl]-2-fluorobenzyl}-N-methyl-beta-alaninate

Step 1: tert-butyl N-(4-cyano-2-fluorobenzyl)-N-methyl-beta-alaninate

The title compound was prepared following the general procedure 10,starting from 4-cyano-2-fluorobenzyl bromide (Fluorochem, 2.0 g, 9.3mmol) and tert-butyl N-methyl-beta-alaninate (1.8 g, 11.2 mmol, preparedas described in Biorg. Med. Chem. (11) 2003, 3083-3099). It was isolatedas a yellow oil (2.4 g, 87%). ¹H NMR (CDCl₃, 300 MHz) δ 7.82 (dd,J=10.0, 1.6 Hz, 1H), 7.69 (dd, J=7.8, 1.6 Hz, 1H), 7.60 (m, 1H), 3.58(s, 2H), 2.61 (t, J=6.9 Hz, 2H), 2.38 (t, J=6.9 Hz, 2H), 2.14 (s, 3H),1.38 (s, 9H). LC/MS (Method B): 293.0 (M+H)⁺. HPLC (Method A) Rt 2.44min (Purity: 88.5%).

Step 2: tert-butylN-{4-[amino(hydroxyimino)methyl]-2-fluorobenzyl}-N-methyl-beta-alaninate

The title compound was prepared following the general procedure 2,starting from tert-butylN-(4-cyano-2-fluorobenzyl)-N-methyl-beta-alaninate (1.4 g, 4.8 mmol). Itwas obtained as a yellow oil (2.4 g, 90%). ¹H NMR (DMSO-d₆, 300 MHz) δ9.74 (br s, 1H), 7.49 (dd, J=8.0, 1.6 Hz, 1H), 7.42 (dd, J=11.6, 1.6 Hz,1H), 7.37 (m, 1H), 5.87 (br s, 2H), 3.51 (s, 2H), 2.60 (t, J=7.0 Hz,2H), 2.38 (t, J=7.0 Hz, 2H), 2.13 (s, 3H), 1.38 (s, 9H). LC/MS (MethodB): 326.0 (M+H)⁺. HPLC (Method A) Rt 1.57 min (Purity: 95.0%).

Intermediate 40: tert-butylN-{4-[amino(hydroxyimino)methyl]benzyl}-N-methyl-beta-alaninate

Step 1: tert-butyl N-(4-cyanobenzyl)-N-methyl-beta-alaninate

The title compound was prepared following the general procedure 10,starting from 4-cyanobenzyl bromide (1.5 g, 7.7 mmol) and tert-butylN-methyl-beta-alaninate (1.5 g, 9.2 mmol, prepared as described inBiorg. Med. Chem. (11) 2003, 3083-3099). It was isolated as a yellow oil(1.9 g, 89%). ¹H NMR (DMSO-d₆, 300 MHz) δ 7.78 (d, J=8.3 Hz, 2H), 7.49(d, J=8.3 Hz, 2H), 3.55 (s, 2H), 2.60 (t, J=6.9 Hz, 2H), 2.38 (t, J=6.9Hz, 2H), 2.11 (s, 3H), 1.39 (s, 9H). LC/MS (Method B): 275.0 (M+H)⁺.HPLC (Method A) Rt 2.40 min (Purity: 82.5%).

Step 2: tert-butylN-{4-[amino(hydroxyimino)methyl]benzyl}-N-methyl-beta-alaninate

The title compound was prepared following the general procedure 2,starting from tert-butyl N-(4-cyanobenzyl)-N-methyl-beta-alaninate (1.9g; 6.8 mmol). It was isolated as a yellow oil (1.6 g, 77%). ¹H NMR(DMSO-d₆, 300 MHz) δ 9.57 (br s, 1H), 7.61 (d, J=8.2 Hz, 2H), 7.27 (d,J=8.2 Hz, 2H), 5.77 (s, 2H), 3.51-3.45 (m, 2H), 2.59 (t, J=7.0 Hz, 2H),2.38 (t, J=7.0 Hz, 2H), 2.10 (s, 3H), 1.40 (s, 9H). LC/MS (Method B):308.0 (M+H)⁺. HPLC (Method A) Rt 1.52 min (Purity: 82.3%).

Intermediate 41: tert-butylN-(3-{5-[4-bromo-3-(methoxymethyl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate

Step 1: 4-bromo-3-(methoxymethyl)benzoic acid

To a solution of methyl 4-bromo-3-(methoxymethyl)benzoate (Intermediate1, Step 2, 7.0 g, 27.0 mmol) in EtOH (210 mL) was added a 5 N aqueoussolution of NaOH (16 mL, 80.0 mmol). The resulting mixture was heated at60° C. for 1 hour. The reaction mixture was cooled to RT andconcentrated under vacuum. The residue was taken up with water andwashed with EtOAc. The aqueous layer was then acidified with a 1Naqueous solution of HCl and extracted with EtOAc. The organic layer wasdried (MgSO₄) and concentrated under vacuum to give the title compoundas a yellow solid (5.81 g, 87%). ¹H NMR (DMSO-d₆, 300 MHz) 13.19 (br s,1H), 8.00 (m, 1H), 7.77 (m, 2H), 4.49 (s, 2H), 3.40 (s, 3H). LC/MS(Method B): 245.0 (M−H)⁻. HPLC (Method A) Rt 3.63 min (purity: 97.4%).

Step 2: tert-butylN-(3-{5-[4-bromo-3-(methoxymethyl)phenyl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinate

Oxalyl chloride (7.8 mL, 91.8 mmol) and DMF (100 μL) were added into asolution of 4-bromo-3-(methoxymethyl)benzoic acid (15.0 g, 61.2 mmol) inanhydrous toluene (225 mL). The resulting mixture was stirred at RT for2 hours, and then concentrated under vacuum to give the acyl chloridederivative as a yellow oil. This acyl chloride was taken up withanhydrous toluene (150 mL) and added drop wise into a solution ofIntermediate 21 (18.0 g, 61.2 mmol) in toluene (75 mL) and pyridine (75mL). The resulting mixture was stirred at RT for 2 hours, and thenheated at reflux for 18 hours. The reaction mixture was concentratedunder vacuum. The residue was taken up with EtOAc (400 mL) and washedwith water (150 mL), a saturated aqueous solution of NaHCO₃ (2×150 mL)and brine (150 mL). The organic layer was dried (MgSO₄) and concentratedunder vacuum to give a yellow solid which was washed with MeOH to givethe title compound as a beige powder. HPLC (Method A) Rt 4.8 min(purity: 96.3%). ¹H NMR (DMSO-d₆, 300 MHz) δ 8.23 (s, 1H), 8.07-7.91 (m,4H), 7.54 (m, 2H), 4.57 (s, 2H), 3.74 (s, 2H), 3.47 (s, 3H), 3.22 (s,2H), 2.29 (s, 3H), 1.45 (s, 9H).

Intermediate 42: methyl4-[amino(hydroxyimino)methyl]-2,5-difluorobenzoate

Step 1: methyl 4-cyano-2,5-difluorobenzoate

Methyl 2,4,5-trifluorobenzoate (DSL Chemicals, 950 mg, 5 mmol), sodiumcyanide (306 mg, 6.25 mmol) and tetrabutylammonium bromide (2.01 g, 6.25mmol) were dissolved in DMF (10 mL) and the resulting mixture was heatedat 60° C. overnight. Additional amount of sodium cyanide (306 mg, 6.25mmol) was added and the mixture was stirred at 60° C. for 24 additionalhours. The reaction mixture was diluted with EtOAc and washed with brineseveral times. The organic layer was dried over MgSO₄ and concentratedunder vacuum. The residue was purified by flash chromatography (silica,EtOAc/cHex) to give the title compound. ¹H NMR (DMSO-d₆, 300 MHz) δ 7.77(dd, J=8.4, 5.5 Hz, 1H), 7.42 (dd, J=8.9, 5.0 Hz, 1H), 3.96 (s, 3H).LC/MS (Method B): 463.2 (M−H)⁻, 465.2 (M+H)⁺. HPLC (Method A) Rt 3.63min (Purity: 99.9%).

Step 2: methyl 4-[amino(hydroxyimino)methyl]-2,5-difluorobenzoate

The title compound was prepared following the general protocol 1,starting from methyl 4-cyano-2,5-difluorobenzoate. It was obtained as awhite solid (304 mg, 98%). LC/MS (Method B): 229.0 (M−H)⁻, 231.0 (M+H)⁺.HPLC (Method A) Rt 1.08 min (Purity: 98.7%).

Intermediate 43: tert-butylN-{3-[amino(hydroxyimino)methyl]-5-chlorobenzyl}-N-methylglycinate

Step 1: 3-(bromomethyl)-5-chlorobenzonitrile

The title compound was prepared following general procedure 13, startingfrom 3-chloro-5-methylbenzonitrile (FluoroChem Ltd). It was isolated asa yellow powder. LC/MS (Method B): 295.1 (M+H)⁺. HPLC (Method A) Rt 4.61min (Purity: 96.8%).

Step 2: tert-butyl N-(3-chloro-5-cyanobenzyl)-methylglycinate

The title compound was prepared following general procedure 10, startingfrom 3-(bromomethyl)-5-chlorobenzonitrile and sarcosine tert-butyl esterhydrochloride. It was isolated as a yellow oil (570 mg, 78%). LC/MS(Method B): 295.1 (M+H)⁺.

Step 3: tert-butylN-{3-[amino(hydroxyimino)methyl]-5-chlorobenzyl}-N-methylglycinate

The title compound was prepared following general procedure 2, startingfrom tert-butyl N-(3-chloro-5-cyanobenzyl)-N-methylglycinate. It wasisolated as a yellowish oil (550 mg, 88%). LC/MS (Method B): 326.2(M−H)⁻.

Intermediate 44: tert-butylN-{4-[amino(hydroxyimino)methyl]-2-methoxybenzyl}-N-methylglycinate

Step 1: tert-butyl N-(4-cyano-2-methoxybenzyl)-N-methylglycinate

The title compound was prepared following the general procedure 10,starting from 4-cyano-2-methoxybenzyl bromide (Carbocore) and sarcosinetert-butyl ester hydrochloride. It was isolated as a colorless oil. ¹HNMR (DMSO-d₆, 300 MHz) δ 7.53 (d, J=7.7 Hz, 1H), 7.45-7.40 (m, 2H), 3.83(s, 3H), 3.69 (s, 2H), 3.20 (s, 2H), 2.27 (s, 3H), 1.42 (s, 9H). LC/MS(Method B): 291.0 (M+H)⁺. HPLC (Method A) Rt 2.52 min (Purity: 93.5%).

Step 2: tert-butylN-{4-[amino(hydroxyimino)methyl]-2-methoxybenzyl}-N-methylglycinate

The title compound was prepared following the general procedure 2,starting from tert-butyl N-(4-cyano-2-methoxybenzyl)-N-methylglycinate(1.0 g, 3.4 mmol). It was isolated as a yellow oil (860 mg, 77%). ¹H NMR(DMSO-d₆, 300 MHz) δ 9.58 (s, 1H), 7.31-7.23 (m, 3H), 5.81 (s, 2H), 3.79(s, 3H), 3.63 (s, 2H), 3.14 (s, 2H), 2.26 (s, 3H), 1.42 (s, 9H). LC/MS(Method B): 324.0 (M+H)⁺. HPLC (Method A) Rt 1.49 min (Purity: 88.6%).

Intermediate 45: tert-butylN-{4-[amino(hydroxyimino)methyl]-2-fluorobenzyl}-N-(tert-butoxycarbonyl)glycinate

Step 1: tert-butyl N-(4-cyano-2-fluorobenzyl)glycinate

The title compound was prepared following the general procedure 10,starting from 4-cyano-2-fluorobenzyl bromide (Fluorochem) and tert-butylglycinate. It was isolated as a yellow oil (1.8 g, 74%). ¹H NMR(DMSO-d₆, 300 MHz) δ 7.82-7.77 (m, 1H), 7.71-7.63 (m, 2H), 3.82 (s, 2H),3.22 (s, 2H), 2.58 (br s, 1H), 1.40 (s, 9H). LC/MS (Method B): 265.0(M+H)⁺. HPLC (Method A) Rt 2.23 min (Purity: 98.0%).

Step 2: tert-butylN-(tert-butoxycarbonyl)-N-(4-cyano-2-fluorobenzyl)glycinate

To a solution of tert-butyl N-(4-cyano-2-fluorobenzyl)glycinate (1.8 g,6.8 mmol) and di-tert-butyl dicarbonate (1.6 g, 7.5 mmol) in DCM (36 mL)was added N-ethyldiisopropylamine (1.7 mL, 10.2 mmol). The resultingmixture was stirred at RT for 3.5 hours. The reaction mixture wasdiluted with DCM and washed with a saturated aqueous solution of NaHCO₃(2×) and brine. The organic layer was dried (MgSO₄) and concentrated togive the title compound as a yellowish oil (2.1 g, 85%). ¹H NMR(DMSO-d₆, 300 MHz) δ 7.84 (m, 1H), 7.71 (m, 1H), 7.58 (m, 1H), 4.52-4.48(m, 2H), 3.95-3.87 (m, 2H), 1.48-1.31 (m, 18H). LC/MS (Method B): 365.1(M+H)⁺. HPLC (Method A) Rt 5.20 min (Purity: 97.1%).

Step 3: tert-butylN-{4-[amino(hydroxyimino)methyl]-2-fluorobenzyl}-N-(tert-butoxycarbonyl)glycinate

The title compound was prepared following the general procedure 2,starting from tert-butylN-(tert-butoxycarbonyl)-N-(4-cyano-2-fluorobenzyl)glycinate (2.1 g, 5.8mmol). It was isolated as a yellow oil (1.7 g, 74%). ¹H NMR (DMSO-d₆,300 MHz) δ 9.76 (s, 1H), 7.54-7.31 (m, 3H), 5.88 (s, 2H), 4.44 (s, 2H),3.89-3.81 (m, 2H), 1.40-1.35 (m, 18H). LC/MS (Method B): 396.2 (M−H)⁻,398.0 (M+H)⁺. HPLC (Method A) Rt 3.39 min (Purity: 95.6%).

Intermediate 47:2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-carboxylic acid

Step 1: methyl2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-carboxylate

A mixture of methyl 4-bromo-3-(methoxymethyl)benzoate (Intermediate 1Step 2, 8.0 g, 30.8 mmol), 2-(trifluoromethyl)phenylboronic acid (8.8 g,46.3 mmol), cesium fluoride (14.0 g, 92.4 mmol), palladium acetate (0.69g, 3.1 mmol) and 2-dicyclohexylphosphino-2,6-dimethoxybiphenyl (3.16 g,7.7 mmol) was prepared in a mixture of dioxane (80 mL) and water (80 mL)under nitrogen atmosphere and heated at 100° C. for 12 hours. Thereaction mixture was concentrated under vacuum. The residue was dilutedwith EtOAc (200 mL), and then washed with water (100 mL) and brine (100mL). The organic layer was dried (Na₂SO₄) and concentrated under vacuum.After purification by flash chromatography (silica, pet ether/EtOAc),the title compound was obtained as a pale yellow solid (8.0 g, 88%). ¹HNMR (CDCl₃, 400 MHz) δ 8.22 (1H, s), 7.99 (1H, d, J=6.5 Hz), 7.78 (1H,d, J=7.7 Hz), 7.57 (2H, m), 7.27 (2H, m), 4.19-4.10 (2H, m), 3.95 (3H,s), 3.26 (3H, s).

Step 2: 2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-carboxylic acid

Lithium hydroxide monohydrate (2.29 g, 54.7 mmol) was added into asolution of methyl2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-carboxylate (8.0 g,27.3 mmol) in THF (100 mL), MeOH (30 mL) and water (30 mL). Theresulting mixture was stirred at RT for 12 hours, and then the reactionmixture was concentrated under vacuum. The remaining aqueous layer wasacidified with a concentrated aqueous solution of HCl and extracted withEtOAc. Then the organic layer was washed with water and brine, and thendried (Na₂SO₄) and concentrated under vacuum. The resulting solid wassuspended in Et₂O (30 mL), filtered and dried under vacuum to afford ofthe title compound as a pale yellow solid (6.2 g, 74%). LC/MS (MethodB): 309.0 (M−H)⁻. ¹H NMR (DMSO-d₆, 400 MHz) δ 13.08 (1H, s), 8.06 (1H,s), 7.87 (2H, m), 7.73 (1H, m), 7.65 (1H, m), 7.35 (1H, d, J=7.5 Hz),7.27 (1H, d, J=7.9 Hz), 4.11-4.03 (2H, m), 3.15 (3H, s).

Intermediate 50: 2-methyl-2′-(trifluoromethyl)biphenyl-4-carboxylic acid

A mixture of methyl 4-bromo-3-methylbenzoate (20.0 g, 87.3 mmol),2-(trifluoromethyl)benzeneboronic acid (24.9 g, 131.0 mmol), potassiumcarbonate (24.1 g, 174.6 mmol) and bis(tricyclohexylphosphine)palladium(II) dichloride (64.5 mg, 0.09 mmol) was prepared in dioxane (200 mL)and water (50 mL) under N₂ atmosphere. The mixture was heated at 100° C.for 3 hours. A 5N aqueous solution of NaOH (100 mL) was added and thereaction mixture was stirred at 100° C. for one additional hour. Thereaction mixture was cooled at RT and the aqueous layer was removed. Theorganic layer was filtered through a Celite pad, concentrated until 75ml under reduced pressure, diluted with water (125 ml) and washed withMTBE (2×200 mL). The aqueous layer was acidified with a 5N aqueoussolution of HCl (25 ml, PH˜1) and extracted with MTBE (2×100 ml). Theorganic layers were combined, dried (Na₂SO₄) and filtered through aCelite pad. The solution was concentrated until 100 mL, then heptane wasadded (200 mL). The mixture was concentrated until 100 mL. Theprecipitate was filtered off and rinsed twice with heptane, then driedunder reduced pressure to give the title compound as a white powder(22.5 g, 92%). HPLC (Method A), Rt 4.4 min (purity: 100%). LC/MS (MethodB): 279.0 (M−H)⁻. ¹H NMR (DMSO-d₆, 300 MHz) δ 13.00 (s, 1H), 7.87 (m,2H), 7.80 (dd, J=7.9, 1.6 Hz, 1H), 7.75 (m, 1H), 7.64 (m, 1H), 7.34 (d,J=7.6 Hz, 1H), 7.23 (d, J=7.9 Hz, 1H), 2.02 (s, 3H).

Intermediate 51: 2′-ethyl-2-(trifluoromethyl)biphenyl-4-carboxylic acid

Step 1: methyl 4-bromo-3-(trifluoromethyl)benzoate

To a suspension of 4-bromo-3-(trifluoromethyl)benzoic acid (APACPharmaceuticals, 20 g, 74.35 mmol) in MeOH (400 ml) at 0° C. was addedthionyl chloride (21.6 ml, 297 mmol) dropwise. The reaction mixture wasstirred at RT for 24 hours. The solvent was concentrated under vacuumand the crude residue was diluted with EtOAc (250 mL). The organic layerwas washed with a saturated NaHCO₃ solution (100 mL), water (100 mL),brine (100 mL), dried over MgSO₄, filtered off and concentrated undervacuum to afford the title compound as a yellow solid (19.13 g, 91%).HPLC (Method A) Rt 5.12 min (Purity: 99.2%).

Step 2: methyl 2′-ethyl-2-(trifluoromethyl)biphenyl-4-carboxylate

A mixture of methyl 4-bromo-3-(trifluoromethyl)benzoate (3.0 g, 10.6mmol), 2-ethylphenylboronic acid (2.38 g, 15.9 mmol), cesium fluoride(4.83 g, 31.8 mmol), palladium acetate (48 mg, 0.21 mmol) and2-dicyclohexylphosphino-2′,6′-dimethoxybiphenyl (261 mg, 0.64 mmol) wasprepared in dioxane (30 mL) and water (15 mL), and then heated at 90° C.for 2.5 hours. The reaction mixture was diluted with MTBE (150 mL) andwashed with water (50 mL) and brine (50 mL). The aqueous layers wereextracted with MTBE (75 mL). The organic layers were combined, dried(MgSO₄) and concentrated under vacuum. After purification by flashchromatography (silica, heptane/EtOAc), the title compound was obtainedas a colorless oil (3.05 g, 93%). HPLC (Method A) Rt 5.6 min (Purity:97.3%).

Step 3: 2′-ethyl-2-(trifluoromethyl)biphenyl-4-carboxylic acid

A 5N aqueous solution of NaOH (3 mL, 15 mmol) was added into a solutionof methyl 2′-ethyl-2-(trifluoromethyl)biphenyl-4-carboxylate (3.05 g,9.89 mmol) in EtOH (30 mL) and the resulting mixture was heated at 60°C. for 45 minutes. The reaction mixture was concentrated under vacuum.The residue was taken up with water (50 mL) and a 5N aqueous solution ofHCl (5 mL), and then extracted with MTBE (2×100 mL). The organic layerswere washed with brine (50 mL), combined, dried (MgSO₄) and the solventwas removed under reduced pressure to give the title compound as a whitepowder (2.71 g, 93%). HPLC (Method A) Rt 4.8 min (Purity: 97.3%). LC/MS(Method B): 293.1 (M−H)⁻. ¹H NMR (DMSO-d₆, 300 MHz) δ 13.54 (s, 1H),8.29 (d, J=1.5 Hz, 1H), 8.22 (dd, J=7.9, 1.5 Hz, 1H), 7.50 (d, J=7.9 Hz,1H), 7.42-7.32 (m, 2H), 7.24 (m, 1H), 7.10 (d, J=7.5 Hz, 1H), 2.39-2.13(m, 2H), 1.00 (t, J=7.6 Hz, 3H).

Intermediate 52: 2-ethoxy-2′-(trifluoromethyl)biphenyl-4-carboxylic acid

Step 1: methyl 2-hydroxy-2′-(trifluoromethyl)biphenyl-4-carboxylate

A mixture of methyl 4-bromo-3-hydroxybenzoate (Combi-Blocks CA-4189,4.00 g, 17.3 mmol), 2-(trifluoromethyl)phenylboronic acid (3.95 g, 20.8mmol), cesium fluoride (7.90 g, 52 mmol), palladium(II) acetate (78 mg,0.35 mmol) and 2-dicyclohexylphosphino-2′,6′-dimethoxybiphenyl (426 mg,1.05 mmol) was prepared in dioxane (40 mL) and water (20 mL) under N₂atmosphere. The reaction mixture was heated at 90° C. After 90 minutes,additional amounts of 2-(trifluoromethyl)phenylboronic acid (1.90 g, 10mmol), palladium(II) acetate (78 mg, 0.35 mmol) and2-dicyclohexylphosphino-2′,6′-dimethoxybiphenyl (426 mg, 1.05 mmol) wereadded. After 45 additional minutes, additional amounts of2-(trifluoromethyl)phenylboronic acid (2.7 g, 14.2 mmol), palladium(II)acetate (78 mg, 0.35 mmol) and2-dicyclohexylphosphino-2′,6′-dimethoxybiphenyl (426 mg, 1.05 mmol) wereadded. After 15 hours, the reaction mixture was cooled to RT, dilutedwith MTBE (200 mL), and then washed with water (30 mL) and brine (50mL). The aqueous layers were extracted with MTBE (100 mL). The organiclayers were combined, dried over MgSO₄ and concentrated under reducedpressure. After purification by flash chromatography (silica,EtOAc/heptane), the title compound was obtained as a yellow oil (5.03 gcontaining 12% w/w of DCM). HPLC (Method A), Rt 3.6 min (purity: 98.8%).LC/MS (Method B): 297.0 (M+H)⁺. 295.0 (M−H)⁻.

Step 2: methyl 2-ethoxy-2′-(trifluoromethyl)biphenyl-4-carboxylate

A mixture of methyl 2-hydroxy-2′-(trifluoromethyl)biphenyl-4-carboxylate(5.03 g), anhydrous potassium carbonate (7.05 g, 51 mmol) andbromoethane (6.35 mL, 85 mmol) was prepared in anhydrous ACN (75 mL) andheated at 50° C. for 15 hours. The reaction mixture was cooled at RT,filtered and concentrated under reduced pressure. After purification byflash chromatography (silica, EtOAc/heptane), the title was obtained asa colorless oil (4.45 g, 79% over 2 steps). HPLC (Method A), Rt 5.3 min(purity: 99.0%). LC/MS (Method B): 325.0 (M+H)⁺.

Step 3: 2-ethoxy-2′-(trifluoromethyl)biphenyl-4-carboxylic acid

A 5 N aqueous solution of NaOH (4.1 mL, 20.5 mmol) was added into asolution of methyl 2-ethoxy-2′-(trifluoromethyl)biphenyl-4-carboxylate(4.45 g, 13.7 mmol) in EtOH (45 mL). The reaction mixture was heated at60° C. for 30 minutes, and then concentrated under reduced pressure. Theresidue was taken up with water (50 mL) and a 5N aqueous solution of HCl(7 ml), and then extracted with MTBE (2×100 mL). The organic layers werewashed with brine (50 mL), dried over MgSO₄ and concentrated underreduced pressure to give a colorless oil. After precipitation frompentane, the title compound was obtained as a white powder. HPLC (MethodA), Rt 4.3 min (purity: 99.7%). LC/MS (Method B): M⁻ (ESI): 309.0.

Intermediate 54: 2-chloro-2′-(trifluoromethyl)biphenyl-4-carboxylic acid

Step 1: methyl 2-chloro-2′-(trifluoromethyl)biphenyl-4-carboxylate

A mixture of methyl 4-borono-3-chlorobenzoate (Combi-Blocks BB-3530, 1.5g, 7.0 mmol), 2-bromobenzotrifluoride (1.9 g, 8.4 mmol), cesium fluoride(3.2 g; 21.0 mmol) and bis(triphenylphosphine)palladium(II) chloride (98mg, 0.14 mmol) was prepared in dioxane (15 mL) and water (7.5 mL) undernitrogen atmosphere, and then heated at 90° C. for 30 minutes. Thereaction mixture was diluted with MTBE (30 mL) and washed with water andbrine. The aqueous layers were extracted with MTBE (30 mL). The combinedorganic layers were dried (MgSO4) and concentrated under vacuum. Aftertwo purifications by flash chromatography (silica, heptane/DCM, thenheptane/EtOAc), the title compound was obtained as a yellow oil. HPLC(Method A) Rt 5.3 min (Purity: 99.7%).

Step 2: 2-chloro-2′-(trifluoromethyl)biphenyl-4-carboxylic acid

A 5N aqueous solution of NaOH (4 mL, 20 mmol) was added into a solutionof methyl 2-chloro-2′-(trifluoromethyl)biphenyl-4-carboxylate (1.07 g,3.40 mmol) in MeOH (16 mL) and stirred at RT for 30 minutes. Thereaction mixture was concentrated under vacuum. The residue was taken upwith EtOAc and washed with a 1N aqueous solution of HCl (3×) and brine,and then dried (MgSO₄) and concentrated under vacuum to give the titlecompound as a white powder (950 mg, 93%). HPLC (Method A) Rt 4.9 min(Purity: 99.4%). LC/MS (Method B): 299.1 (M−H)⁻. ¹H NMR (DMSO-d₆, 300MHz) δ 13.48 (s, 1H), 8.03 (d, J=1.6 Hz, 1H), 7.94 (dd, J=7.9, 1.6 Hz,1H), 7.89 (d, J=7.8 Hz, 1H), 7.78 (m, 1H), 7.69 (m, 1H), 7.49 (d, J=7.9Hz, 1H), 7.41 (d, J=7.4 Hz, 1H).

Intermediate 56: 2′-fluoro-2-(trifluoromethyl)biphenyl-4-carboxylic acid

A mixture of 4-bromo-3-(trifluoromethyl)benzoic acid (15.0 g, 55.7mmol), 2-fluorophenylboronic acid (9.4 g, 66.9 mmol), cesium fluoride(25.4 g, 167 mmol), 2-dicyclohexylphosphino-2′,6′-dimethoxybiphenyl (5.7g, 13.9 mmol) and palladium(II) acetate (1.25 g, 5.6 mmol) was preparedin dioxane (150 mL) and water (150 mL) under N₂ atmosphere. The reactionmixture was purged with vacuum for 5 minutes, and then the reactionmixture was degassed with N₂ at RT for 10 min. The reaction mixture washeated at 110° C. for 3 hours. The reaction mixture was cooled to RT andwas filtered through a pad of Celite. The filtrate was concentratedunder vacuum affording a yellow solid. After purification by flashchromatography (silica, DCM containing 1% of AcOH), the title compoundwas obtained as a pale yellow powder. HPLC (Method A), Rt 4.3 min(purity: 99.9%). LC/MS (Method B): 283.0 (M−H)⁻. ¹H NMR (DMSO-d₆, 300MHz) δ 13.60 (s, 1H), 8.31 (d, J=1.4 Hz, 1H), 8.25 (dd, J=8.0, 1.4 Hz,1H), 7.60 (d, J=8.0 Hz, 1H), 7.52 (m, 1H), 7.38-7.26 (m, 3H).

Intermediate 58: tert-butylN-{2-fluoro-3-[(hydroxyamino)(imino)methyl]benzyl}-N-methylglycinate

Step 1: 3-(bromomethyl)-2-fluorobenzonitrile

The title compound was prepared following the general procedure 13,starting from 2-fluoro-3-methylbenzonitrile (Matrix Scientific). It wasobtained as an orange solid. HPLC (Method A) Rt 3.72 (Purity: 99.3%).

Step 2: tert-butylN-{2-fluoro-3-[(hydroxyamino)(imino)methyl]benzyl}-N-methylglycinate

Tert-butyl N-(3-cyano-2-fluorobenzyl)-N-methylglycinate was madefollowing the general procedure 10, starting from3-(bromomethyl)-2-fluorobenzonitrile and sarcosine tert-butyl esterhydrochloride. It was isolated as an oil (1.05 g, 88%). It was submittedto the general procedure 2, affording the title compound as a yellow oil(1 g, 85%). HPLC (Method A) Rt 1.09 (Purity: 92%).

Intermediate 59: tert-butylN-{3-[amino(hydroxyimino)methyl]benzyl}-N-isopropyl-beta-alaninate

Step 1: 3-[(isopropylamino)methyl]benzonitrile

The title compound was prepared following general procedure 10, startingfrom 3-(bromomethyl)benzonitrile and isopropyl amine. It was isolated asa yellow liquid (5.7 g, 92%). ¹H NMR (DMSO-d₆, 400 MHz) δ 7.73 (s, 1H),7.67 (d, J=7.7 Hz, 1H), 7.63 (d, J=7.8 Hz, 1H), 7.49 (m, 1H), 3.56 (s,2H), 2.75 (m, 1H), 0.83 (d, J=8.8 Hz, 6H).

Step 2: tert-butyl N-(3-cyanobenzyl)-N-isopropyl-beta-alaninate

A mixture of 3-[(isopropylamino)methyl]benzonitrile (5.7 g, 32.7 mmol),tert-butyl acrylate (4.78 ml, 32.7 mmol) and DBU (5.0 ml, 32.7 mmol) washeated neat at 80° C. for 24 hours. The reaction mixture was dilutedwith water and extracted with DCM. The organic layer was dried oversodium sulfate and concentrated under reduced pressure. Afterpurification by chromatography (silica, pet ether/EtOAc), the titlecompound was obtained as a colorless oil. ¹H NMR (DMSO-d₆, 400 MHz) δ7.73 (s, 1H,), 7.67 (d, J=7.7 Hz, 1H), 7.63 (d, J=7.8 Hz, 1H), 7.49 (t,J=7.7 Hz, 1H), 3.56 (s, 2H), 2.75 (m, 1H), 2.62 (t, J=6.5 Hz, 2H), 2.25(t, J=6.4 Hz, 2H), 1.39 (s, 9H), 0.83 (d, J=8.8 Hz, 6H).

Step 3: tert-butylN-{3-[amino(hydroxyimino)methyl]benzyl}-N-isopropyl-beta-alaninate

The title compound was prepared following general procedure 2, startingfrom tert-butyl N-(3-cyanobenzyl)-N-isopropyl-beta-alaninate. It wasobtained as a colorless oil (4.8 g, 90%). ¹H NMR (DMSO-d₆, 400 MHz) δ9.56 (s, 1H), 7.59 (s, 1H), 7.50 (d, J=7.6 Hz, 1H), 7.34 (d, J=7.6 Hz,1H), 7.27 (m, 1H), 5.73 (s, 2H), 3.51 (s, 2H), 2.80 (m, 1H), 2.62 (t,J=6.8 Hz, 2H), 2.27 (t, J=6.8 Hz, 2H), 1.37 (s, 9H), 0.94 (d, J=6.6 Hz,6H). LC/MS (Method A): 336.3 (M+H)⁺. HPLC (Method A) Rt 2.19 min(Purity: 97.4%).

Intermediate 60: tert-butylN-{5-[amino(hydroxyimino)methyl]-2-chlorobenzyl}-N-methylglycinate

Step 1: 3-(bromomethyl)-4-chlorobenzonitrile

The title compound was prepared following the general procedure 13,starting from 4-chloro-3-methylbenzonitrile (TransWorld Chemicals) andwas isolated as a pale yellow powder (1.8 g, 59%). HPLC (Method A) Rt4.05 (Purity: 97%).

Step 2: tert-butyl N-(2-chloro-5-cyanobenzyl)-N-methylglycinate

The title compound was prepared following general procedure 10, startingfrom 3-(bromomethyl)-4-chlorobenzonitrile and sarcosine tert-butyl esterhydrochloride. It was obtained as a colorless oil (1.9 g, 86%). LC/MS(Method B): 295.1 (M+H)⁺.

Step 3: tert-butylN-{5-[amino(hydroxyimino)methyl]-2-chlorobenzyl}-N-methylglycinate

The title compound was prepared following general procedure 1, startingfrom tert-butyl N-(2-chloro-5-cyanobenzyl)-N-methylglycinate and wasisolated as a colorless oil (1.55 g, 73%). LC/MS (Method B): 328.1(M+H)⁺.

Intermediate 61: tert-butyl4-[{4-[amino(hydroxyimino)methyl]benzyl}(methyl)amino]butanoate

Step 1: tert-butyl 4-[(4-cyanobenzyl)(methyl)amino]butanoate

The title compound was prepared following general procedure 10, startingfrom 4-cyanobenzyl bromide and tert-butyl 4-(methylamino)butanoatehydrochloride (Watanabe). It was obtained as a yellow oil (586 mg,quantitative). LC/MS (Method B): 289.2 (M+H)⁺. HPLC (Method A) Rt 3.02(Purity: 91.8%).

Step 2: tert-butyl4-[{4-[amino(hydroxyimino)methyl]benzyl}(methyl)amino]butanoate

The title compound was prepared following general procedure 2, startingfrom tert-butyl 4-[(4-cyanobenzyl)(methyl)amino]butanoate and wasisolated as an off-white solid (468 mg, 73%). HPLC (Method A) Rt 2.1(Purity: 98.7%). LC/MS (Method B): 322.2 (M+H)⁺.

Intermediate 64: 3,5-difluoro-N′-hydroxy-4-(hydroxymethyl)benzimidamide

Step 1: 3,5-difluoro-4-(hydroxymethyl)benzonitrile

A mixture of (4-bromo-2,6-difluorophenyl)methanol (1.12 g, 5.0 mmol),Pd(PPh₃)₄ (323 mg, 0.28 mmol) and zinc cyanide (587 mg, 5.0 mmol) wasprepared in DMF (7 mL) under nitrogen atmosphere and heated at 90° C.for 18 hours. The mixture was diluted with DCM and water. The organicphase was passed through a hydrophobic frit and the solvent evaporatedin vacuo. The residue was purified by flash chromatography (silica,iso-hexane/EtOAc) to afford the title compound. ¹H NMR (CDCl₃, 400 MHz)δ 7.28-7.21 (2H, m), 4.83 (2H, d, J=6.7 Hz), 1.95 (1H, t, J=6.7 Hz).

Step 2: 3,5-difluoro-N′-hydroxy-4-(hydroxymethyl)benzimidamide

The title compound was prepared following the general procedure 1,starting from 3,5-difluoro-4-(hydroxymethyl)benzonitrile. It wasobtained as an off-white solid (512 mg, 84%). ¹H NMR (DMSO-d₆, 400 MHz)δ 9.95 (1H, s), 7.39 (2H, d, J=8.5 Hz), 6.00 (2H, br s), 5.30 (1H, t,J=5.6 Hz), 4.52 (2H, d, J=5.6 Hz).

Intermediate 65: tert-butyl2-((3-chloro-4-(N′-hydroxycarbamimidoyl)benzyl)(methyl)amino)acetate

Step 1: tert-butyl 2-((4-cyano-3-chlorobenzyl)(methyl)amino)acetate

A solution of 4-methyl-2-chlorobenzonitrile (1.51 g, 10 mmol),N-bromosuccinimide (2.14 g, 12.0 mmol) and AIBN (33 mg, 0.20 mmol) inACN (40 mL) was heated under reflux for 18 hours. The reaction mixturewas diluted with EtOAc and washed with a saturated aqueous solution ofNa₂CO₃ and brine. The organic layer was dried (MgSO₄) and concentratedunder vacuum. The residue was treated with sarcosine tert-butyl esterhydrochloride (1.44 g, 7.92 mmol) and K₂CO₃ (2.74 g, 19.8 mmol) in ACN(10 mL). The mixture was heated at 100° C. for 2 hours. The solvent wasevaporated under vacuum. The residue was dissolved in a mixture of DCMand water and then poured through a hydrophobic frit. The solvent wasevaporated under vacuum. The residue was purified by flashchromatography (silica, iso-hexane/Et₂O) to afford the title compound(1.95 g, quantitative). ¹H NMR (CDCl₃, 400 MHz) δ 7.61 (1H, d, J=8.0Hz), 7.57 (1H, s), 7.36 (1H, m), 3.73 (2H, s), 3.21 (2H, s), 2.36 (3H,s), 1.48 (9H, s).

Step 2: tert-butyl2-((3-chloro-4-(N′-hydroxycarbamimidoyl)benzyl)(methyl)amino)acetate

A solution of tert-butyl2-((4-cyano-3-chlorobenzyl)(methyl)amino)acetate (1.95 g, 6.60 mmol) and50% aqueous hydroxylamine (2.02 mL) in ethanol (10 mL) was heated at 80°C. for 5 hours. The solvent was evaporated in vacuo. The residue waspartitioned between DCM and water. The organic phase was poured througha hydrophobic frit and evaporated in vacuo to afford the title compound(2.16 g, quantitative). ¹H NMR (CDCl₃, 400 MHz) δ 7.48-7.42 (2H, m),7.28 (1H, m), 4.97 (2H, s), 3.69 (2H, s), 3.18 (2H, s), 2.37 (3H, s),1.48 (9H, s).

Intermediate 66: N′-hydroxy-4-(hydroxymethyl)-3-methylbenzimidamide

Step 1: (4-bromo-2-methylphenyl)methanol

To a solution of 4-bromo-2-methylbenzoic acid (1.0 g, 4.65 mmol) in Et₂O(15 mL) was added lithium aluminumhydride (370 mg, 10.23 mmol) and themixture was stirred for 4 hours. The reaction mixture was diluted withwater (100 mL) and extracted with EtOAc (4×100 mL). The combined organicfractions were dried (MgSO₄) and concentrated in vacuo. The residue waspurified by flash chromatography (silica, petrol ether/EtOAc) to givethe title compound as a colorless gum (752 mg, 80%). ¹H NMR (CDCl₃, 400MHz) δ 7.35-7.30 (2H, m), 7.24 (1H, m), 4.65 (2H, s), 2.32 (3H, s).

Step 2: 4-(hydroxymethyl)-3-methylbenzonitrile

A mixture of (4-bromo-2-methylphenyl)methanol (603 mg, 3.0 mmol),Pd₂(dba)₃ (110 mg, 0.12 mmol), S-Phos (99 mg, 0.24 mmol) and zinccyanide (421 mg, 3.6 mmol) was prepared in a mixture of DMF/water (99:1,5 mL) under nitrogen atmosphere and heated at 130° C. for 30 minutesunder microwave irradiation. The suspension was filtered through aCelite pad and the filter-cake washed with EtOAc. The filtrate waswashed with water and brine. The organic phase was passed through ahydrophobic frit and the solvent evaporated in vacuo. The residue waspurified by flash chromatography (silica, petrol ether/EtOAc) to givethe title compound. ¹H NMR (CDCl₃, 400 MHz) δ 7.58-7.48 (2H, m), 7.44(1H, s), 4.75 (2H, d, J=5.5 Hz), 2.33 (3H, s), 1.85-1.78 (1H, m).

Step 3: N′-hydroxy-4-(hydroxymethyl)-3-methylbenzimidamide

The amidoxime was prepared following the procedure 1, but starting from4-(hydroxymethyl)-3-methylbenzonitrile. It was obtained as a white solid(741 mg, 71%). ¹H NMR (DMSO-d₆, 400 MHz) δ 9.51 (1H, s), 7.46 (2H, m),7.34 (1H, d, J=7.7 Hz), 5.72 (2H, s), 5.09 (1H, s), 4.50 (2H, s), 2.26(3H, s).

Intermediate 67: tert-butylN-{4-[amino(hydroxyimino)methyl]-2-ethylbenzyl}-N-methylglycinate

Step 1: tert-butyl N-(4-cyano-2-vinylbenzyl)-N-methylglycinate

A mixture of tert-butyl N-(2-bromo-4-cyanobenzyl)-N-methylglycinate(Intermediate 33 Step 1, 658 mg, 1.94 mmol), vinyl boronic acid pinacolester (597 mg, 3.88 mmol), K₂CO₃ (536 mg, 3.88 mmol) andtetrakis-triphenylphosphine palladium(0) (224 mg, 0.19 mmol) wasprepared in a mixture of dioxane/water (5:1; 6 mL) and heated at 100° C.for 18 hours. The reaction mixture was diluted with DCM/water andseparated. The organic phase was passed through a hydrophobic frit andthe solvents evaporated in vacuo. The residue was purified by flashchromatography (silica, iso-hexane/EtOAc) to afford the title compound(508 mg, 92%). ¹H NMR (CDCl₃, 400 MHz) δ 7.77 (1H, s), 7.58-7.43 (2H,m), 7.13 (1H, dd, J=17.4, 11.0 Hz), 5.69 (1H, d, J=17.4 Hz), 5.42 (1H,d, J=11.0 Hz), 3.80 (2H, s), 3.18 (2H, s), 2.36 (3H, s), 1.50 (9H, s).

Step 2: tert-butyl N-(4-cyano-2-ethylbenzyl)-N-methylglycinate

A mixture of tert-butyl N-(4-cyano-2-vinylbenzyl)-N-methylglycinate (500mg, 1.75 mmol) and 10% palladium on carbon (50 mg) in methanol (5 mL)were stirred under an atmosphere of hydrogen for 18 hours. Thesuspension was filtered through a Celite pad and the filter-cake washedwith DCM. The filtrate was evaporated in vacuo to afford the titlecompound (474 mg, 94%).

Step 3: tert-butylN-{4-[amino(hydroxyimino)methyl]-2-ethylbenzyl}-N-methylglycinate

A solution of tert-butyl N-(4-cyano-2-ethylbenzyl)-N-methylglycinate(570 mg, 1.98 mmol) and 50% aqueous hydroxylamine (0.6 mL, 10 mmol) inethanol (10 mL) was heated at 80° C. for 18 hours. The solvent wasevaporated in vacuo. The residue was partitioned between DCM and water.The organic phase was passed through a hydrophobic frit and evaporatedin vacuo to afford the title compound (658 mg, quantitative). ¹H NMR(CDCl₃, 400 MHz) δ 7.46 (1H, s), 7.39 (2H, s), 4.85 (2H, br s), 3.73(2H, s), 3.16 (2H, s), 2.73 (2H, q, J=7.6 Hz), 2.37 (3H, s), 1.47 (9H,s), 1.25-1.19 (3H, m).

Intermediate 68: N′-hydroxy-3-(hydroxyethyl)benzenecarboximidamide

The title compound was prepared following the general procedure 1, butstarting from 3-(hydroxyethyl)benzonitrile (912 mg, 6.20 mmol). It wasobtained as a white solid (1.05 g, 94%). ¹H NMR (DMSO-d₆, 400 MHz) δ9.59 (1H, s), 7.55 (1H, s), 7.51 (1H, d, J=7.7 Hz), 7.33-7.21 (2H, m),5.77 (2H, s), 4.66 (1H, t, J=5.2 Hz), 3.65 (2H, m), 2.77 (2H, t, J=7.1Hz). LC/MS (Method A): 181 (M+H)⁺. HPLC (Method J) Rt=8.26 min (Purity:99.9%).

Intermediate 69: N′-hydroxy-4-(hydroxyethyl)benzenecarboximidamide

The title compound was prepared following the general procedure 1, butstarting from 4-(hydroxyethyl)benzonitrile (2.43 g, 16.5 mmol). It wasobtained as a white solid (2.95 g, 99%). ¹H NMR (DMSO-d₆, 400 MHz) δ9.58 (1H, s), 7.60 (2H, d, J=7.9 Hz), 7.24 (2H, d, J=7.9 Hz), 5.75 (2H,s), 4.72 (1H, m), 3.64 (2H, m), 2.76 (2H, m). LC/MS (Method A): 181(M+H)⁺. HPLC (Method J) Rt=8.15 min (Purity: 98.7%).

Intermediate 70: N′-hydroxy-3-(hydroxymethyl)benzimidamide

The title compound was prepared following the general procedure 1, butstarting from 3-(hydroxymethyl)benzonitrile (8.43 g, 63.4 mmol). It wasobtained as a white solid (9.15 g, 86%). ¹H NMR (DMSO-d₆, 400 MHz) δ9.63 (1H, s), 7.67 (1H, s), 7.56 (1H, m), 7.35 (2H, m), 5.82 (2H, s),5.27 (1H, t, J=5.7 Hz), 4.54 (2H, d, J=5.7 Hz).

Intermediate 71: tert-butylN-{4-[amino(hydroxyimino)methyl]benzyl}-N-(2-methoxyethyl)glycinate

Step 1: 4-{[(2-methoxyethyl)amino]methyl}benzonitrile

2-Methoxyethylamine (2.2 mL, 25.5 mmol) was added into a solution of4-cyanobenzyl bromide (1.0 g, 5.1 mmol) in ACN (10 mL) and the resultingmixture was stirred at RT for 1 hour. The reaction mixture wasconcentrated under vacuum. The crude product was dissolved in EtOAc, andthen washed with a saturated aqueous solution of NaHCO₃ and brine, dried(MgSO₄) and concentrated under vacuum to give the title compound (958mg, 99%). LC/MS (Method B): 191.0 (M+H)⁺. HPLC (Method A) Rt 1.44 min(Purity: 93.4%).

Step 2: tert-butyl N-(4-cyanobenzyl)-N-(2-methoxyethyl)glycinate

Tert-butyl bromoacetate (1.64 mL, 11.1 mmol) was added into a mixture of4-{[(2-methoxyethyl)amino]methyl}benzonitrile (958 mg, 5.04 mmol) andK₂CO₃ (3.06 g, 22.2 mmol) in ACN (20 mL). The resulting mixture wasstirred at RT overnight. The reaction mixture was diluted with EtOAc,and then washed with water and brine, dried (MgSO₄) and concentratedunder vacuum. The residue was purified by flash chromatography (silica,cHex/EtOAc) to give the title compound as a colorless oil. LC/MS (MethodB): 305.1 (M+H)⁺. HPLC (Method A) Rt 3.09 min (Purity: 91.1%).

Step 3: tert-butylN-{4-[amino(hydroxyimino)methyl]benzyl}-N-(2-methoxyethyl)glycinate

The title compound was prepared according the general procedure 1,starting from tert-butyl N-(4-cyanobenzyl)-N-(2-methoxyethyl)glycinateand was isolated as an oil. ¹H NMR (DMSO-d₆, 400 MHz) δ 9.58 (br s, 1H),7.61 (d, J=8.0 Hz, 1H), 7.29 (d, J=8.0 Hz, 1H), 5.78 (s, 2H), 3.77 (s,2H), 3.44-3.36 (m, 2H), 3.25 (s, 2H), 3.20 (s, 3H), 2.79-2.71 (m, 2H),1.42 (s, 9H). LC/MS (Method B): 338.2 (M+H)⁺.

Intermediate 72: N′-hydroxy-4-(hydroxymethyl)benzenecarboximidamide

The title compound was prepared according the general procedure 1,starting from 4-(hydroxymethyl)benzonitrile. It was obtained as a whitesolid (13.1 g, 95%). ¹H NMR (DMSO-d₆, 400 MHz) δ 9.58 (1H, s), 7.66 (2H,m), 7.34 (2H, d, J=8.1 Hz), 5.79 (2H, s), 5.23 (1H, t, J=5.6 Hz), 4.54(2H, d, J=5.6 Hz).

Intermediate 73: N′-hydroxy-4-(hydroxymethyl)-2-methylbenzimidamide

Step 1: 4-(hydroxymethyl)-2-methylbenzonitrile

Lithium borohydride (33 mg, 1.54 mmol) was added into a solution ofmethyl 4-cyano-3-methylbenzoate (245 mg, 1.40 mmol) in THF (3 mL) andthe resulting mixture was stirred at RT for 4 hours. The reactionmixture was diluted with water (10 mL) and extracted with EtOAc (4×20mL). The combined organic fractions were dried (MgSO₄) and concentratedunder vacuum. The residue was purified by flash chromatography (silica,petrol ether/EtOAc) to give the title compound as a colorless gum (195mg, 95%). ¹H NMR (CDCl₃, 400 MHz) δ 7.58 (1H, d, J=7.9 Hz), 7.33 (1H,s), 7.26 (1H, m), 4.74 (2H, s), 2.55 (3H, s), 1.95 (1H, s). LC/MS(Method A): 144 (M+H)⁺. HPLC (Method F) Rt 3.95 min (Purity: 95.3%).

Step 2: N′-hydroxy-4-(hydroxymethyl)-2-methylbenzimidamide

The title compound was prepared following the general procedure 1,starting from 4-(hydroxymethyl)-2-methylbenzonitrile. It was obtained asa white solid. ¹H NMR (DMSO-d₆, 400 MHz) δ 9.29 (1H, s), 7.24 (1H, d,J=7.7 Hz), 7.18-7.09 (2H, m), 5.68 (2H, s), 5.17 (1H, t, J=5.7 Hz), 4.48(2H, d, J=5.7 Hz), 2.35 (3H, s). LC/MS (Method A): 181 (M+H)⁺. HPLC(Method F) Rt 1.10 min (Purity: 91.3%).

Intermediate 74: tert-butyl2-((4-(N′-hydroxycarbamimidoyl)-2-(trifluoromethyl)benzyl)(methyl)amino)acetate

Step 1: tert-butyl2-((4-cyano-2-(trifluoromethyl)benzyl)(methyl)amino)acetate

A solution of 4-methyl-3-(trifluoromethyl)benzonitrile (1.85 g, 10mmol), N-bromosuccinimide (2.14 g, 12.0 mmol) and AIBN (0.033 g, 0.2mmol) in ACN (40 mL) was heated under reflux for 18 hours. The reactionmixture was diluted with EtOAc and water. The suspension was filteredand the filtrate was separated. The organic phase was washed with brine,dried (MgSO₄) and concentrated under vacuum. The residue was purified byflash chromatography (silica, iso-hexane/EtOAc). The material wastreated with sarcosine tert-butyl ester hydrochloride (0.516 g, 2.84mmol) and K₂CO₃ (0.980 g, 7.1 mmol) in ACN (10 mL). The mixture washeated at 70° C. for 18 hours. The solvent was evaporated under vacuum.The residue was dissolved in a mixture of DCM and water and then pouredthrough a hydrophobic frit. The solvent was evaporated under vacuum. Theresidue was purified by flash chromatography (silica, iso-hexane/EtOAc)to afford the title compound (0.543 g, 70%). ¹H NMR (CDCl₃, 400 MHz) δ δ8.15 (1H, d, J=8.2 Hz), 7.90 (1H, s), 7.83 (1H, d, J=8.2 Hz), 3.90 (2H,s), 3.26 (2H, s), 2.37 (3H, s), 1.48 (9H, s).

Step 2: tert-butyl2-((4-(N′-hydroxycarbamimidoyl)-2-(trifluoromethyl)benzyl)(methyl)amino)acetate

A solution of tert-butyl2-((4-cyano-2-(trifluoromethyl)benzyl)(methyl)amino)acetate (0.541 g,1.65 mmol) and 50% aqueous hydroxylamine (0.50 mL) in ethanol (2 mL) washeated at 75° C. for 2 hours. The solvent was evaporated in vacuo. Theresidue was partitioned between DCM and water. The organic phase waspassed through a hydrophobic frit and evaporated in vacuo to afford thetitle compound (0.460 g, 77%). ¹H NMR (CDCl₃, 400 MHz,) δ 7.97 (1H, d,J=8.1 Hz), 7.89 (1H, s), 7.78 (1H, d, J=8.1 Hz), 4.86 (2H, br s), 3.86(2H, s), 3.23 (2H, s), 2.37 (3H, s), 1.48 (9H, s).

Intermediate 75: tert-butyl2-((2-chloro-4-(N′-hydroxycarbamimidoyl)benzyl)(methyl)amino)acetate

Step 1: tert-butyl 2-((4-cyano-2-chlorobenzyl)(methyl)amino)acetate

A solution of 4-methyl-3-chlorobenzonitrile (4.69 g, 31 mmol),N-bromosuccinimide (6.04 g, 34.2 mmol) and benzoyl peroxide (0.750 g,3.10 mmol) in ACN (20 mL) was heated under reflux for 18 hours. Thereaction mixture was diluted with EtOAc and washed with a saturatedaqueous solution of Na₂CO₃ and brine. The organic phase was dried(MgSO4) and concentrated under vacuum. A portion of the residue (1.0 g,4.34 mmol) was treated with sarcosine tert-butyl ester hydrochloride(1.57 g, 8.69 mmol) and K₂CO₃ (2.39 g, 17.4 mmol) in dioxane (5 mL). Theresulting mixture was heated at 100° C. for 2 hours, and thenconcentrated under vacuum. The residue was dissolved in a mixture of DCMand water, and then passed through a hydrophobic frit. The solvent wasevaporated under vacuum to afford the title compound (0.952 g, 74%).

Step 2: tert-butyl2-((2-chloro-4-(N′-hydroxycarbamimidoyl)benzyl)(methyl)amino)acetate

A solution of tert-butyl2-((4-cyano-2-chlorobenzyl)(methyl)amino)acetate (1.25 g, 4.25 mmol) and50% aqueous hydroxylamine (1.30 mL) in ethanol (10 mL) was heated at 80°C. for 5 hours. The solvent was evaporated under vacuum. The residue waspartitioned between DCM and water. The organic phase was passed througha hydrophobic frit and concentrated under vacuum to afford the titlecompound (1.34 g, 76%). ¹H NMR (CDCl₃, 400 MHz) δ 7.63 (1H, d, J=1.8Hz), 7.59 (1H, d, J=8.0 Hz), 7.50 (1H, dd, J=8.0, 1.8 Hz), 4.84 (2H, s),3.83 (2H, s), 3.25 (2H, s), 2.42 (3H, s), 1.48 (9H, s).

Intermediate 76: tert-butyl2-((2,6-difluoro-4-(N′-hydroxycarbamimidoyl)benzyl)(methyl)amino)acetate

Step 1: tert-butyl 2-((4-cyano-2,6-difluorobenzyl)(methyl)amino)acetate

To a degassed solution of 3,5-difluoro-4-methylbenzonitrile (1.53 g, 10mmol) and AIBN (0.033 g; 0.20 mmol) in ACN (40 mL) was addedN-bromo-succinimide (2.14 g, 12.0 mmol). The reaction mixture was heatedat 90° C. for 24 hours. The solvent was evaporated in vacuo and theresidue partitioned between water and DCM. The organic phase was passedthrough a hydrophobic frit and the solvent evaporated in vacuo. Theresidue was dissolved in CAN, and then sarcosine t-butyl esterhydrochloride (1.09 g, 6.0 mmol) and K₂CO₃ (2.07 g, 15 mmol) added. Thereaction mixture was stirred at 70° C. for 18 hours. The suspension wasfiltered and the filtrate evaporated in vacuo. The residue was purifiedby flash chromatography (silica, iso-hexane/EtOAc) to afford the titlecompound. ¹H NMR (CDCl₃, 400 MHz) δ 7.22 (2H, d, J=5.9 Hz), 3.94 (2H,s), 3.21 (2H, s), 2.41 (3H, s), 1.48 (9H, s).

Step 2: tert-butyl2-((2,6-difluoro-4-(N′-hydroxycarbamimidoyl)benzyl)(methyl)amino)acetate

A solution of tert-butyl2-((4-cyano-2,6-difluorobenzyl)(methyl)amino)acetate (0.281 g, 0.95mmol) and 50% aqueous hydroxylamine (0.32 mL) in ethanol (1.3 mL) washeated at 75° C. for 18 hours. The solvent was evaporated in vacuo. Theresidue was partitioned between EtOAc and water, and then the organicphase was passed through a hydrophobic frit. The solvent was evaporatedin vacuo to afford the title compound. ¹H NMR (CDCl₃, 400 MHz) δ7.22-7.13 (2H, m), 4.81 (2H, br s), 3.91 (2H, s), 3.20 (2H, s), 2.40(3H, s), 1.48 (9H, s).

Example 14N-methyl-N-(3-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycine

Tert-butylN-methyl-N-(3-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycinatewas prepared following the general procedure 7 starting fromIntermediate 21 and Intermediate 5. It was hydrolyzed followingprocedure 8, affording the title compound as a white powder. ¹H NMR(DMSO-d₆, 300 MHz) δ 8.55 (s, 1H), 8.50 (d, J=8.0 Hz, 1H), 8.36 (s, 1H),8.23 (d, J=7.8 Hz, 1H), 7.83 (d, J=7.8 Hz, 1H), 7.78-7.63 (m, 2H),7.42-7.12 (m, 3H), 7.17 (d, J=7.4 Hz, 1H), 4.51 (s, 2H), 4.12 (s, 2H),2.81 (s, 3H), 2.02 (s, 3H). HPLC (Method A); Rt 4.20 min (Purity:99.48%). CHN analysis: [C₂₆H₂₂O₃F₃N₃—HCl] Calculated: C, 60.29%, H,4.48%, N, 8.11%. Found: C, 60.08%, H, 4.42%, N, 7.91%.

Example 49N-(3-{5-[2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(3-{5-[2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinatewas prepared following the general procedure 3 starting fromIntermediate 21 and Intermediate 47. After purification by flashchromatography (silica, heptane/EtOAc), the ester derivative washydrolyzed following the general procedure 8. After purification byprecipitation from a mixture of ACN and water, the title compound wasobtained as a white powder (876 mg, 70% over 2 steps). HPLC (Method A),Rt 4.1 min (purity: 99.6%). LC/MS (Method B): 510.1 (M−H)⁻, 512.0(M+H)⁺. Melting point: 207° C. Elemental analysis: [C₂₇H₂₄N₃O₄F₃—HCl]Corrected: C, 59.18%, H, 4.60%, N, 7.67%, Cl 6.47%. Found: C, 58.96%, H,4.52%, N, 7.78%, Cl 6.69%. ¹H NMR (DMSO-d₆, 300 MHz) δ 8.35 (s, 1H),8.33 (d, J=1.8 Hz, 1H), 8.22 (d, J=7.8 Hz, 1H), 8.17 (dd, J=8.0, 1.8 Hz,1H), 7.92 (d, J=7.5 Hz, 1H), 7.84-7.68 (m, 4H), 7.48 (d, J=8.0 Hz, 1H),7.43 (d, J=7.3 Hz, 1H), 4.51 (s, 2H), 4.22 (d, J=13.0 Hz, 1H), 4.13 (d,J=13.0 Hz, 1H), 4.12 (s, 2H), 3.25 (s, 3H), 2.82 (s, 3H).

Example 50N-(3-{5-[4′-fluoro-2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(3-{5-[4′-fluoro-2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinatewas prepared following the general procedure 15 starting fromIntermediate 41 and 4-fluoro-2-(trifluoromethyl)benzeneboronic acid(Fluorochem 33830). After purification by flash chromatography (silica,cHex/EtOAc), the ester derivative was hydrolyzed following the generalprocedure 8. After purification by precipitation from ACN, the titlecompound was obtained as a white powder. HPLC (Method A), Rt 3.2 min(purity: 97.3%). LC/MS (Method B): 528.1 (M−H)⁻, 530.0 (M+H)⁺. ¹H NMR(DMSO-d₆, 300 MHz) δ 8.33 (m, 2H), 8.22 (d, J=7.8 Hz, 1H), 8.18 (dd,J=7.9, 1.6 Hz, 1H), 7.86-7.79 (m, 2H), 7.75-7.64 (m, 2H), 7.51 (m, 2H),4.50 (s, 2H), 4.22 (d, J=13.0 Hz, 1H), 4.15 (d, J=13.0 Hz, 1H), 4.11 (s,2H), 3.25 (s, 3H), 2.81 (s, 3H).

Example 64N-(3-fluoro-5-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(3-fluoro-5-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinatewas prepared following the general procedure 3 starting fromIntermediate 5 and Intermediate 24. It was hydrolyzed following generalprocedure 8 to afford the title compound as a white powder. ¹H NMR(DMSO-d₆, 300 MHz) δ 8.56 (m, 1H), 8.51 (dd, J=7.6 Hz, 1H), 8.20 (m,1H), 8.01 (m, 1H), 7.71 (m, 2H), 7.43-7.34 (m, 2H), 7.17 (m, 1H), 4.45(brs, 2H), 4.07 (brs, 2H), 2.79 (s, 3H), 2.03 (s, 3H). LC/MS (Method B):500.2 (M+H)⁺, 498.2 (M−H)⁻. HPLC (Method A) Rt 4.92 min (Purity: 99.8%).

Example 69N-ethyl-N-(3-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-beta-alanine,hydrochloride salt

Tert-butylN-ethyl-N-(3-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-beta-alaninatewas prepared following the general procedure 3 starting fromIntermediate 5 and Intermediate 37. It was deprotected following generalprocedure 8 to afford the title compound as a white powder. ¹H NMR(DMSO-d₆, 300 MHz) δ 12.75 (brs, 1H), 10.30 (brs, 1H), 8.55 (m, 1H),8.51 (d, J=8.0 Hz, 1H), 8.23 (d, J=7.9 Hz, 1H), 7.90 (m, 1H), 7.72 (m,2H), 7.43-7.33 (m, 2H) 7.29 (m, 1H), 7.17 (m, 1H), 4.50 (brs, 2H), 3.29(m, 2H), 3.13 (m, 2H), 2.83 (m, 2H), 2.03 (s, 3H), 1.28 (t, J=7.2 Hz,3H). LC/MS (Method B): 510.2 (M+H)⁺, 508.2 (M−H)⁻. HPLC (Method A) Rt4.90 min (Purity: 99.6%).

Example 70N-(2-fluoro-4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(2-fluoro-4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinatewas prepared following the general procedure 4 starting fromIntermediate 5 and Intermediate 32. It was deprotected following thegeneral procedure 8, affording the title compound as a white powder. ¹HNMR (DMSO-d₆, 300 MHz) δ 8.56 (d, J=1.3 Hz, 1H), 8.51 (dd, J=7.9, 1.5Hz, 1H), 8.09 (dd, J=8.0, 1.5 Hz, 1H), 8.02 (dd, J=10.2, 1.3 Hz, 1H),7.91 (t, J=7.7 Hz, 1H), 7.69 (d, J=8.0 Hz, 1H), 7.42-7.26 (m, 3H), 7.17(d, J=7.5 Hz, 1H), 4.50 (s, 2H), 4.13 (s, 2H), 2.81 (s, 3H), 2.03 (s,3H). LC/MS (Method B): 498.1 (M−H)⁻, 500.0 (M+H)⁺. HPLC (Method A) Rt4.35 min (Purity: 99.9%). Elemental analysis: [C₂₆H₂₁N₃O₃F₄—HCl—H₂O]Corrected: C, 56.38%, H, 4.37%, N, 7.59%, Cl 6.40%. Found: C, 56.18%, H,4.29%, N, 7.51%, Cl 6.12. Melting point: 183° C.

Example 71N-(2-fluoro-4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methyl-beta-alanine,hydrochloride salt

Tert-butylN-(2-fluoro-4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methyl-beta-alaninatewas prepared following the general procedure 4 starting fromintermediate 5 and intermediate 39. It was deprotected following thegeneral procedure 8 affording the title compound as a white powder. ¹HNMR (DMSO-d₆, 300 MHz) δ 8.56 (d, J=1.3 Hz, 1H), 8.51 (dd, J=8.0, 1.4Hz, 1H), 8.09 (dd, J=8.0, 1.5 Hz, 1H), 8.06-7.97 (m, 2H), 7.68 (d, J=8.1Hz, 1H), 7.42-7.26 (m, 3H), 7.17 (d, J=7.5 Hz, 1H), 4.50 (s, 2H), 3.45(br s, 2H), 2.90 (t, J=7.5 Hz, 2H), 2.75 (s, 3H), 2.03 (s, 3H). LC/MS(Method A): 512.1 (M−H)⁻, 514.0 (M+H)⁺. HPLC (Method A) Rt 4.36 min(Purity: 99.6%).

Example 81N-methyl-N-(4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycine,hydrochloride salt

Tert-butylN-methyl-N-(4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycinatewas prepared following the general procedure 4 starting fromintermediate 5 and intermediate 31. It was hydrolyzed following thegeneral procedure 8 affording the title compound as a white powder. ¹HNMR (DMSO-d₆, 300 MHz) δ 8.55 (s, 1H), 8.51 (d, J=8.0 Hz, 1H), 8.23 (d,J=8.1 Hz, 2H), 7.80 (d, J=8.1 Hz, 2H), 7.68 (d, J=7.9 Hz, 1H), 7.42-7.26(m, 3H), 7.17 (d, J=7.3 Hz, 1H), 4.46 (s, 2H), 4.10 (s, 2H), 2.80 (s,3H), 2.03 (s, 3H). LC/MS (Method B): 480.1 (M−H)⁻, 482.0 (M+H)⁺. HPLC(Method A) Rt 4.76 min (Purity: 100.0%).

Example 89N-(4-{5-[2′-fluoro-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(4-{5-[2′-fluoro-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinatewas prepared following the general procedure 4 starting fromIntermediate 56 and Intermediate 31. It was hydrolyzed following thegeneral procedure 8 affording the title compound as a white powder. ¹HNMR (DMSO-d₆, 300 MHz) δ 8.58-8.52 (m, 2H), 8.24 (d, J=8.3 Hz, 2H),7.83-7.77 (m, 3H), 7.62-7.54 (m, 1H), 7.46-7.32 (m, 3H), 4.46 (s, 2H),4.10 (s, 2H), 2.81 (s, 3H). LC/MS (Method B): 484.1 (M−H)⁻, 486.0(M+H)⁺. HPLC (Method A) Rt 4.08 min (Purity: 99.9%).

Example 95N-(2-fluoro-4-{5-[2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(2-fluoro-4-{5-[2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinatewas prepared following the general procedure 3 starting fromIntermediate 47 and Intermediate 32. It was hydrolyzed following generalprocedure 8 to afford the title compound as a white powder. LC/MS(Method B): 530.0 (M+H)⁺, 528.1 (m-H)⁻. HPLC (Method A) Rt 4.14 min(Purity: 98.9%).

Example 97N-(4-{5-[2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(4-{5-[2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinatewas prepared following the general procedure 3 starting fromIntermediate 47 and Intermediate 31. It was hydrolyzed following generalprocedure 8 to afford the title compound as an off-white powder. LC/MS(Method B): 512.0 (M+H)⁺, 510.0 (M−H)⁻. HPLC (Method A) Rt 4.04 min(Purity: 98.8%).

Example 100N-methyl-N-(4-{5-[2-methyl-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycine,hydrochloride salt

Tert-butylN-methyl-N-(4-{5-[2-methyl-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycinatewas prepared following the general procedure 3 starting fromIntermediate 50 and Intermediate 31. It was deprotected following thegeneral procedure 8 affording the title compound as a white powder (433mg, 75%). ¹H NMR (DMSO-d₆, 300 MHz) δ 8.25-8.16 (m, 3H), 8.12-8.05 (m,1H), 7.91 (d, J=7.2 Hz, 1H), 7.84-7.76 (m, 3H), 7.74-7.65 (m, 1H),7.47-7.37 (m, 2H), 4.47 (s, 2H), 4.11 (s, 2H), 2.81 (s, 3H), 2.12 (s,3H). LC/MS (Method B): 480.1 (M−H)⁻, 482.0 (M+H)⁺. HPLC (Method A) Rt4.18 min (Purity 99.8%). Elemental analysis: [C₂₆H₂₂N₃O₃F₃—HCl]Calculated: C, 60.29%, H, 4.48%, N, 8.11%, Cl 6.85%. Found: C, 60.14%,H, 4.36%, N, 8.12%, Cl 6.81%.

Example 101N-methyl-N-(3-{5-[2-methyl-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycine,hydrochloride salt

Tert-butylN-isopropyl-N-(3-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-beta-alaninatewas prepared following the general procedure 3 starting fromIntermediate 50 and Intermediate 21. It was hydrolyzed following generalprocedure 8 to afford the title compound as a white powder. ¹H NMR(DMSO-d₆, 300 MHz) δ 8.33 (brs, 1H), 8.21 (m, 1H), 8.16 (m, 1H), 8.07(m, 1H), 8.01 (m, 1H), 7.84-7.65 (m, 4H), 7.42 (m, 2H), 4.50 (brs, 2H),4.12 (brs, 2H), 2.82 (s, 3H), 2.12 (s, 3H). LC/MS (Method B): 482.0(M+H)⁺, 480.2 (M−H)⁻. HPLC (Method A) Rt 4.17 min (Purity: 99.7%).

Example 1032-(methyl(3-methyl-4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)amino)aceticacid

The title compound was prepared following the procedure described inExample 102, but using Intermediate 5 in Step 1. It was isolated as awhite solid. ¹H NMR (DMSO-d₆, 400 MHz) δ 8.60-8.54 (1H, m), 8.56-8.49(1H, m), 8.07 (1H, d, J=7.7 Hz), 7.69 (1H, d, J=7.9 Hz), 7.46-7.35 (4H,m), 7.35-7.29 (1H, m), 7.21 (1H, d, J=7.5 Hz), 3.78 (2H, s), 3.27 (2H,s), 2.67 (3H, s), 2.34 (3H, s), 2.06 (3H, s). LC/MS (Method A): 496(M+H)⁺. HPLC (Method F) Rt 3.33 min (Purity: 97.8%).

Example 104N-(3-fluoro-5-{5-[2-methyl-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(3-fluoro-5-{5-[2-methyl-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinatewas prepared following the general procedure 4 starting fromIntermediate 50 and Intermediate 24. It was hydrolyzed following thegeneral procedure 8 affording the title compound as a white powder. ¹HNMR (DMSO-d₆, 300 MHz) δ 8.19 (s, 2H), 8.08 (dd, J=7.9, 1.5 Hz, 1H),7.98 (d, J=8.2 Hz, 1H), 7.92 (d, J=7.6 Hz, 1H), 7.84-7.65 (m, 3H),7.47-7.37 (m, 2H), 4.49 (s, 2H), 4.11 (s, 2H), 2.82 (s, 3H), 2.12 (s,3H). LC/MS (Method A): 498.2 (M−H)⁻, 500.1 (M+H)⁺. HPLC (Method A) Rt4.34 min (Purity: 99.7%). Melting point: 207° C.

Example 109N-(4-{5-[2′-ethyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(4-{5-[2′-ethyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinatewas prepared following the general procedure 3 starting fromIntermediate 51 and Intermediate 31. It was hydrolyzed following generalprocedure 8 to afford the title compound as a white powder. ¹H NMR(DMSO-d₆, 300 MHz) δ 8.54 (brs, 1H), 8.50 (m, 1H), 8.23 (d, J=8.3 Hz,2H), 7.79 (d, J=8.3 Hz, 2H), 7.70 (d, J=8.1 Hz, 1H), 7.47-7.37 (m, 2H),7.29 (m, 1H), 7.15 (d, J=7.6 Hz, 1H), 4.45 (brs, 2H), 4.08 (brs, 2H),2.79 (s, 3H), 2.44-2.17 (m, 2H), 1.13 (t, J=7.6 Hz, 3H). LC/MS (MethodB): 496.0 (M+H)⁺, 494.2 (M−H)⁻. HPLC (Method A) Rt 4.44 min (Purity:99.8%).

Example 110N-(4-{5-[2′-ethyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}-2-fluorobenzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(4-{5-[2′-ethyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}-2-fluorobenzyl)-N-methylglycinatewas prepared following the general procedure 3 starting fromIntermediate 51 and Intermediate 32. It was hydrolyzed following generalprocedure 8 to afford the title compound as a white powder. ¹H NMR(DMSO-d₆, 300 MHz) δ 8.54 (d, J=1.4 Hz, 1H), 8.50 (dd, J=8.0, 1.4 Hz,1H), 8.08 (dd, J=7.9, 1.5 Hz, 1H), 8.01 (dd, J=10.2, 1.5 Hz, 1H), 7.93(m, 1H), 7.70 (d, J=8.0 Hz, 1H), 7.41 (m, 2H), 7.28 (m, 1H), 7.15 (d,J=7.6 Hz, 1H), 4.52 (s, 2H), 4.14 (s, 2H), 2.82 (s, 3H), 2.45-2.17 (m,2H), 1.03 (t, J=7.6 Hz, 3H). LC/MS (Method B): 514.0 (M+H)⁺, 512.2(M−H)⁻. HPLC (Method A) Rt 4.50 min (Purity: 99.8%).

Example 111N-(2-chloro-5-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(2-chloro-5-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinatewas prepared following the general procedure 3 starting fromIntermediate 5 and Intermediate 60. It was hydrolyzed following generalprocedure 8 to afford the title compound as a white powder. ¹H NMR(DMSO-d₆, 300 MHz) δ 8.53 (m, 3H), 8.22 (m, 2H), 7.84 (d, J=8.3 Hz, 1H),7.69 (d, J=8.0 Hz, 1H), 7.43-7.33 (m, 2H), 7.29 (m, 1H), 7.17 (m, 1H),4.59 (m, 2H), 4.17 (m, 2H), 2.81 (s, 3H), 2.02 (s, 3H). LC/MS (MethodB): 516.0 (M+H)⁺, 514.1 (M−H)⁻. HPLC (Method A) Rt 4.39 min (Purity:99.6%).

Example 112N-(2-fluoro-3-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(2-fluoro-3-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinatewas prepared following the general procedure 3 starting fromIntermediate 5 and Intermediate 58. It was hydrolyzed following generalprocedure 8 to afford the title compound as a white powder. ¹H NMR(DMSO-d₆, 300 MHz) δ 8.55 (m, 1H), 8.51 (m, 1H), 8.30 (m, 1H), 7.91 (m,1H), 7.68 (d, J=8.0 Hz, 1H), 7.58 (m, 1H), 7.42-7.33 (m, 2H), 7.29 (m,1H), 7.17 (m, 1H), 4.53 (m, 2H), 4.16 (m, 2H), 2.81 (s, 3H), 2.02 (s,3H). LC/MS (Method B): 500.0 (M+H)⁺, 498.1 (M−H)⁻. HPLC (Method A) Rt4.22 min (Purity: 100%).

Example 113(Isopropyl-{4-[5-(2′-methyl-2-trifluoromethyl-biphenyl-4-yl)-[1,2,4]oxadiazol-3-yl]-benzyl}-amino)-aceticacid

Step 1:(4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)phenyl)methanol

To a solution of Intermediate 5 (4.32 g, 15.4 mmol) in MeCN (15 mL) wasadded 1-ethyl-3-dimethyl aminopropylcarbodiimide (4.14 g, 21.6 mmol)followed by Intermediate 72 (2.81 g, 17.0 mmol). The mixture was stirredat RT for 18 hours and then pyridine (5 mL) added. The mixture washeated under microwave irradiation at 150° C. for 30 minutes. Thesolvent was removed in vacuo and the residue partitioned between DCM andH₂O. The layers were separated, the organic layer washed with H₂O andthe mixture passed through a hydrophobic frit. The solvent was removedin vacuo and the residue was recrystallised from EtOAc and petrol togive the title compound as a white solid. ¹H NMR (CDCl₃, 400 MHz) δ 8.63(1H, s), 8.40 (1H, d, J=8.0 Hz), 8.20 (2H, d, J=8.0 Hz), 7.54 (2H, d,J=7.9 Hz), 7.47 (1H, d, J=7.9 Hz), 7.38-7.21 (3H, m), 7.16 (1H, d, J=7.5Hz), 4.81 (2H, d, J=5.7 Hz), 2.14-1.94 (3H, m). LC/MS (Method A): 411(M+H)⁺. HPLC (Method G) Rt 4.50 min (Purity: 97.9%).

Step 2:4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzaldehyde

(4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)phenyl)methanol(5.89 g, 14.4 mmol) was dissolved in dioxane (100 mL) and manganesedioxide (10 g, 116.0 mmol) was added. The mixture was heated at 70° C.overnight and then the solvent was removed in vacuo. The residue wastriturated with a mixture of petrol/diethyl ether to give the titlecompound as a white solid (5.72 g, 97%). ¹H NMR (CDCl₃, 400 MHz) δ 10.13(1H, s), 8.64 (1H, s), 8.43-8.36 (3H, m), 8.06 (2H, d, J=8.0 Hz), 7.49(1H, d, J=8.0 Hz), 7.38-7.21 (3H, m), 7.15 (1H, d, J=7.6 Hz), 2.07 (3H,s). LC/MS (Method A): 409 (M+H)⁺. HPLC (Method G) Rt 4.89 min (Purity:95.7%).

Step 3:(Isopropyl-{4-[5-(2′-methyl-2-trifluoromethyl-biphenyl-4-yl)-[1,2,4]oxadiazol-3-yl]-benzyl}-amino)-aceticacid

Sodium cyanoborohydride (18 mg, 0.28 mmol) was added to a solution of4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzaldehyde(0.25 mmol) and 2-aminoacetic acid (23 mg, 0.25 mmol) in a mixture ofmethanol (3 mL), DCM (3 mL) and acetic acid (38 μl). The mixture wasstirred at room temperature overnight and was filtered through a fritunder positive pressure. To the filtrate was added acetone (67 mg; 1.5mmol) followed by addition of AcOH until the pH was in the range of 3-4(240 mL). To the resulting mixture was added sodium cyanoborohydride (79mg, 1.26 mmol), the mixture stirred for 16 hours, the solvent removed invacuo and the residue purified by reverse phase HPLC to give a whitesolid as the title compound. ¹H NMR (DMSO-d₆, 400 MHz) δ 8.55-8.53 (1H,m), 8.53-8.48 (1H, m), 8.13-8.09 (2H, m), 7.71 (1H, d, J=7.9 Hz), 7.63(2H, d, J=8.0 Hz), 7.37-7.34 (2H, m), 7.28-7.24 (1H, m), 7.15 (1H, d,J=7.5 Hz), 4.02 (2H, s), 3.24-3.22 (3H, m), 2.01 (3H, s), 1.11 (6H, d,J=6.5 Hz). LC/MS (Method A): 510 (M+H)⁺. HPLC (Method F) Rt=3.39 min(Purity: 99.1%).

Example 114N-(2-fluoro-3-{5-[2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(2-fluoro-3-{5-[2-(methoxymethyl)-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinatewas prepared following the general procedure 3 starting fromIntermediate 47 and Intermediate 58. It was hydrolyzed following generalprocedure 8 to afford the title compound as a white powder. ¹H NMR(DMSO-d₆, 300 MHz) δ 8.33 (d, J=1.7 Hz, 1H), 8.28 (m, 1H), 8.18 (dd,J=8.0, 1.7, 1H), 7.90 (m, 2H), 7.79 (m, 1H), 7.71 (m, 1H), 7.57 (m, 1H),7.48 (d, J=8.0 Hz, 1H), 7.43 (d, J=7.3 Hz, 1H), 4.52 (s, 2H), 4.24-4.10(m, 4H), 3.24 (s, 3H), 2.81 (s, 3H). LC/MS (Method B): 530.1 (M+H)⁺,528.2 (M−H)⁻. HPLC (Method A) Rt 4.22 min (Purity: 100%).

Example 115N-(4-{5-[2-ethoxy-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(4-{5-[2-ethoxy-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinatewas prepared following the general procedure 4 starting fromIntermediate 52 and Intermediate 31. It was deprotected following thegeneral procedure 8 affording the title compound as a white powder. ¹HNMR (DMSO-d₆, 300 MHz) δ 8.21 (d, J=8.3 Hz, 2H), 7.85 (dd, J=7.7, 1.5Hz, 2H), 7.81-7.71 (m, 4H), 7.65 (t, J=7.7 Hz, 1H), 7.44 (d, J=7.9 Hz,1H), 7.39 (d, J=7.7 Hz, 1H), 4.44 (s, 2H), 4.25-4.04 (m, 4H), 2.79 (s,3H), 1.18 (t, J=6.9 Hz, 3H). LC/MS (Method B): 510.1 (M−H)⁻, 512.0(M+H)⁺. HPLC (Method A) Rt 4.27 min (Purity: 100%). Melting point: 210°C.

Example 1164-[methyl(4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)amino]butanoicacid, hydrochloride salt

Tert-butyl4-[methyl(4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)amino]butanoatewas prepared following the general procedure 3 starting fromIntermediate 5 and Intermediate 61. It was hydrolyzed following generalprocedure 8 to afford the title compound as a white powder. ¹H NMR(DMSO-d₆, 300 MHz) δ 12.31 (brs, 1H), 10.92 (brs, 1H), 8.53 (brs, 1H),8.50 (d, J=8.5 Hz, 1H), 8.22 (d, J=8.1 Hz, 2H), 7.86 (d, J=8.1 Hz, 2H),7.67 (d, J=7.9 Hz, 1H), 7.42-7.32 (m, 2H), 7.32-7.25 (m, 1H), 7.20-7.12(m, 1H), 4.42 (brs, 2H), 3.08 (brs, 2H), 2.68 (s, 3H), 2.34 (m, 2H),2.02 (s, 3H), 1.98 (m, 2H). LC/MS (Method B): 510.1 (M+H)⁺, 508.2(M−H)⁻. HPLC (Method A) Rt 4.30 min (Purity: 99.6%).

Example 117(Isopropyl-{3-[5-(2′-methyl-2-trifluoromethyl-biphenyl-4-yl)-[1,2,4]oxadiazol-3-yl]-benzyl}-amino)-aceticacid

The title compound was prepared according the procedure described forExample 113, but using Intermediate 70 in Step 1. It was isolated as apale yellow oil. LC/MS (Method B): 508 (M−H)⁻, 510 (M+H)⁺. HPLC (MethodF) Rt 3.41 min (Purity: 99.2%).

Example 123N-(2-fluoro-4-{5-[2-methyl-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine

The title compound was prepared following the general procedure 4starting from intermediate 50 and intermediate 32. It was deprotectedduring the microwave heating affording the title compound by filtrationas a beige powder. ¹H NMR (DMSO-d₆, 300 MHz) δ 12.22 (br s, 1H), 8.17(s, 1H), 8.07 (dd, J=7.9, 1.4 Hz, 1H), 7.96 (dd, J=8.0, 1.6 Hz, 1H),7.91 (d, J=7.8 Hz, 1H), 7.84 (dd, J=10.6, 1.5 Hz, 1H), 7.78 (d, J=7.5Hz, 1H), 7.74-7.65 (m, 2H), 7.46-7.38 (m, 2H), 3.84 (s, 2H), 3.30 (s,2H), 2.33 (s, 3H), 2.12 (s, 3H). LC/MS (Method B): 498.2 (M−H)⁻, 500.1(M+H)⁺. HPLC (Method A) Rt 4.27 min (Purity: 99.6%).

Example 1292-((4-(5-(2-(trifluoromethyl)-2′-methylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl)-2-chlorobenzyl)(methyl)amino)aceticacid

The title compound was prepared following the procedure described inExample 128, but using Intermediate 5 in Step 1. It was obtained as awhite solid. ¹H NMR (DMSO-d₆, 400 MHz) δ 8.54 (1H, s), 8.50 (1H, d,J=8.1 Hz), 8.10-8.04 (2H, m), 7.83 (1H, d, J=7.9 Hz), 7.65 (1H, d, J=8.0Hz), 7.41-7.34 (2H, m), 7.30 (1H, td, J=3.8, 2.2 Hz), 7.17 (1H, d, J=7.6Hz), 3.84 (2H, s), 3.00 (2H, s), 2.31 (3H, s), 2.02 (3H, s). LC/MS(Method A): 516 (M+H)⁺. HPLC (Method F) Rt 3.45 min (Purity: 98.7%).

Example 134N-(4-{5-[2-chloro-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}-2-fluorobenzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(4-{5-[2-chloro-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}-2-fluorobenzyl)-N-methylglycinatewas prepared following the general procedure 3 starting fromintermediate 54 and intermediate 32. It was deprotected following thegeneral procedure 8 affording the title compound as a white powder.LC/MS (Method B): 518.3 (M−H)⁻, 520.2 (M+H)⁺. HPLC (Method A) Rt 4.25min (Purity: 99.8%).

Example 135N-(4-{5-[2-chloro-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl-N-methylglycine,hydrochloride salt

Tert-butylN-(4-{5-[2-chloro-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycinatewas prepared following the general procedure 3 starting fromintermediate 54 and intermediate 31. It was deprotected following thegeneral procedure 8 affording the title compound as a white powder.LC/MS (Method B): 500.2 (M−H)⁻, 502.2 (M+H)⁺. HPLC (Method A) Rt 4.18min (Purity: 99.8%).

Example 136N-(3-{5-[2-chloro-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}-5-fluorobenzyl)-N-methylglycine,hydrochloride salt

Tert-butylN-(3-{5-[2-chloro-2′-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}-5-fluorobenzyl)-N-methylglycinatewas prepared following the general procedure 4 starting fromintermediate 54 and intermediate 24. It was deprotected following thegeneral procedure 8 affording the title compound as a white powder. ¹HNMR (DMSO-d₆, 300 MHz) δ 8.37 (d, J=1.6 Hz, 1H), 8.23 (dd, J=8.0, 1.6Hz, 1H), 8.20 (s, 1H), 8.00 (d, J=8.2 Hz, 1H), 7.93 (d, J=7.7 Hz, 1H),7.87-7.72 (m, 3H), 7.69 (d, J=8.2 Hz, 1H), 7.47 (d, J=7.5 Hz, 1H), 4.51(s, 2H), 4.12 (s, 2H), 2.83 (s, 3H). LC/MS (Method B): 518.2 (M−H)⁻,520.2 (M+H)⁺. HPLC (Method A) Rt 4.82 min (Purity: 99.1%). Meltingpoint: 212° C.

Example 138N-(2-methoxyethyl)-N-(4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycine,hydrochloride salt

Tert-butylN-(2-methoxyethyl)-N-(4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycinatewas prepared following the general procedure 3 with Intermediate 5 andIntermediate 71. It was then hydrolyzed according to general procedure8, affording the title compound as a white solid. ¹H NMR (DMSO-d₆, 300MHz) δ 8.55 (m, 1H), 8.51 (m, 1H), 8.21 (d, J=8.4 Hz, 2H), 7.77 (d,J=7.7 Hz, 2H), 7.68 (d, J=7.7 Hz, 1H), 7.44-7.25 (m, 3H), 7.17 (d, J=7.7Hz, 1H), 4.43 (br s, 2H), 3.95 (br s, 2H), 3.68 (m, 2H), 3.30 (m, 2H),3.27 (s, 3H), 2.03 (s, 3H). LC/MS (Method B): 524.3 (M−H)⁻, 526.3(M+H)⁺. HPLC (Method A) Rt 4.93 min (Purity: 99.5%).

Example 1452-((4-(5-(2-(trifluoromethyl)-2′-methylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl)-3-chlorobenzyl)(methyl)amino)aceticacid, hydrochloride salt

The title compound was prepared according the procedure described forExample 144, but using Intermediate 5 in Step 1. ¹H NMR (DMSO-d₆, 400MHz) δ 8.58 (1H, s), 8.54 (1H, dd, J=8.0, 1.7 Hz), 8.20 (1H, d, J=8.0Hz), 7.96 (1H, s), 7.76 (1H, dd, J=8.0, 1.7 Hz), 7.72 (1H, d, J=8.0 Hz),7.45-7.38 (2H, m), 7.34 (1H, td, J=3.7, 2.3 Hz), 7.21 (1H, d, J=7.6 Hz),4.43 (2H, s), 4.10 (2H, s), 2.82 (3H, s), 2.06 (3H, s). LC/MS (MethodA): 516 (M+H)⁺. HPLC (Method F) Rt 3.34 min (Purity: 99.9%).

Example 1482-((2,6-difluoro-4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)(methyl)amino)aceticacid, hydrochloride salt

Step 1: tert-butyl2-((2,6-difluoro-4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)(methyl)amino)acetate

To a solution of Intermediate 76 (0.148 g, 0.45 mmol) and Intermediate 5(0.107 mg, 0.375 mmol) in ACN (2.5 mL) was added EDC (0.101 g, 0.53mmol). The reaction mixture was stirred at room temperature for 18hours. The reaction mixture was diluted with pyridine (2.5 mL) andheated at 150° C. in the microwave for 30 minutes. The solvent wasremoved in vacuo and the residue dissolved in DCM. The mixture waswashed with water and the organic phase passed through a hydrophobicfrit. The solvent was evaporated in vacuo. The residue was purified byflash chromatography (silica, iso-hexane/EtOAc) to afford the titlecompound. ¹H NMR (CDCl₃, 400 MHz) δ 8.62 (1H, d, J=1.7 Hz), 8.39 (1H,dd, J=8.0, 1.7 Hz), 7.76 (2H, d, J=7.1 Hz), 7.49 (1H, d, J=8.0 Hz),7.38-7.22 (3H, m), 7.15 (1H, d, J=7.6 Hz), 3.99 (2H, s), 3.25 (2H, s),2.45 (3H, s), 2.07 (3H, s), 1.50 (9H, s).

Step 2:2-((2,6-difluoro-4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)(methyl)amino)aceticacid, hydrochloride salt

To tert-butyl2-((2,6-difluoro-4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)(methyl)amino)acetate(0.101 g, 0.18 mmol) was added a 4N solution of HCl in dioxane (4.5 mL)and the reaction mixture stirred at 70° C. for 1 hour. The reactionmixture was allowed to cool and the solvent evaporated in vacuo. Theresidue was triturated with ACN to afford the title compound (0.087 g,87%). ¹H NMR (DMSO-d₆, 400 MHz) δ 8.60 (1H, d, J=1.7 Hz), 8.55 (1H, dd,J=8.0, 1.7 Hz), 7.98 (2H, d, J=7.7 Hz), 7.72 (1H, d, J=8.0 Hz),7.45-7.38 (2H, m), 7.33 (1H, ddd, J=7.6, 6.5, 2.3 Hz), 7.20 (1H, d,J=7.6 Hz), 4.51 (2H, br s), 4.15 (2H, br s), 2.81 (3H, br s), 2.06 (3H,br s). LC/MS (Method A): 518 (M+H)⁺. HPLC (Method F) Rt 3.38 min(Purity: 99.5%).

Example 1502-((2,3-difluoro-4-(5-(2-(trifluoromethyl)-2′-methylbiphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)(methyl)amino)aceticacid, hydrochloride salt

The title compound was prepared according the protocols described forExample 148 and Intermediate 76, but using2,3-difluoro-4-methylbenzonitrile. ¹H NMR (DMSO-d₆, 400 MHz) δ 8.59 (1H,d, J=1.7 Hz), 8.55 (1H, dd, J=8.0, 1.7 Hz), 8.12 (1H, ddd, J=8.2, 6.2,1.6 Hz), 7.78-7.70 (2H, m), 7.45-7.38 (2H, m), 7.33 (1H, ddd, J=7.6,6.5, 2.3 Hz), 7.21 (1H, d, J=7.6 Hz), 4.53 (2H, s), 4.13 (2H, s), 2.84(3H, s), 2.06 (3H, s). LC/MS (Method A): 518 (M+H)⁺. HPLC (Method F) Rt3.34 min (Purity: 99.0%).

Example 1522-((2,3-difluoro-4-(5-(2-methyl-2′-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)(methyl)amino)aceticacid, hydrochloride salt

The title compound was prepared according the protocol described forExample 148, but using Intermediate 50, and the protocol described forIntermediate 76, but using 2,3-difluoro-4-methylbenzonitrile. It wasobtained as a white solid. ¹H NMR (DMSO-d₆, 400 MHz) δ 8.21 (1H, d,J=1.7 Hz), 8.14-8.05 (2H, m), 7.95 (1H, d, J=7.9 Hz), 7.83 (1H, t, J=7.5Hz), 7.76-7.67 (2H, m), 7.46 (2H, m), 4.50 (2H, br s), 4.10 (2H, br s),2.82 (3H, br s), 2.16 (3H, br s). LC/MS (Method A): 518 (M+H)⁺. HPLC(Method F) Rt 3.26 min (Purity: 98.7%).

Example 1532-((3-chloro-4-(5-(2-methyl-2′-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)(methyl)amino)aceticacid, hydrochloride salt

The title compound was prepared according the procedure described inExample 105, but using Intermediate 50 and Intermediate 65. It wasobtained as a white solid. ¹H NMR (DMSO-d₆, 400 MHz) δ 8.20 (1H, d,J=1.7 Hz), 8.16 (1H, d, J=8.0 Hz), 8.11 (1H, dd, J=8.0, 1.8 Hz), 7.95(2H, d, J=8.1 Hz), 7.83 (1H, t, J=7.6 Hz), 7.78-7.69 (2H, m), 7.46 (2H,m), 4.44 (2H, br s), 4.11 (2H, br s), 2.83 (3H, br s), 2.15 (3H, br s).LC/MS (Method A): 516 (M+H)⁺. HPLC (Method F) Rt 3.25 min (Purity:97.5%).

Example 1542-(methyl(2-methyl-4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)amino)aceticacid

(2-methyl-4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)phenyl)methanolwas prepared following the procedure described for Example 113, butusing Intermediate 66, and was isolated as a white solid. It was usedfor the preparation of the title compound, following the proceduredescribed for Example 125. The title compound was isolated as a whitesolid. ¹H NMR (DMSO-d₆, 400 MHz) δ 8.55 (1H, s), 8.51 (1H, d, J=8.1 Hz),7.99-7.92 (2H, m), 7.65 (1H, d, J=8.0 Hz), 7.60 (1H, d, J=7.8 Hz),7.43-7.35 (2H, m), 7.31 (1H, td, J=7.2, 2.1 Hz), 7.18 (1H, d, J=7.6 Hz),3.93 (2H, s), 3.32 (2H, s), 2.48 (3H, s), 2.44 (3H, s), 2.03 (3H, s).LC/MS (Method A): 496 (M+H)⁺. HPLC (Method F) Rt 3.33 min (Purity:99.8%).

Example 1552-((2-ethyl-4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)(methy)amino)aceticacid

Step 1: tert-butylN-(2-ethyl-4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}phenyl)-N-methylglycinate

To a solution of Intermediate 67 (0.222 g, 0.69 mmol) and Intermediate 5(0.232 mg, 0.83 mmol) in ACN (3 mL) was added EDC (0.159 g, 0.83 mmol).The reaction mixture was stirred at room temperature for 18 hours. Thereaction mixture was diluted with pyridine (2 mL) and heated at 150° C.under microwave irradiation for 30 minutes. The solvent was removed invacuo and the residue dissolved in DCM. The mixture was washed withwater and the organic phase passed through a hydrophobic frit. Thesolvent was evaporated in vacuo. The residue was purified by flashchromatography (silica, iso-hexane/EtOAc) affording the title compound.¹H NMR (CDCl₃, 400 MHz) δ 8.64 (1H, d, J=1.2 Hz), 8.42-8.40 (1H, m),8.03 (1H, m), 8.00-7.98 (1H, m), 7.57-7.55 (1H, m), 7.48-7.46 (1H, m),7.36-7.15 (4H, m), 3.81 (2H, s), 3.24 (2H, s), 2.86-2.82 (2H, m), 2.44(3H, s), 2.07 (3H, s), 1.49 (9H, s), 1.32-1.29 (3H, m).

Step 2:2-((2-ethyl-4-(5-(2′-methyl-2-(trifluoromethyl)biphenyl-4-yl)-1,2,4-oxadiazol-3-yl)benzyl)(methyl)amino)aceticacid

To tert-butylN-(2-ethyl-4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}phenyl)-N-methylglycinate(0.109 g; 0.19 mmol) was added a 4N solution of HCl in dioxane (4 mL)and the reaction mixture stirred at room temperature for 18 hours. Thesolvent was evaporated in vacuo and the residue triturated with hotEtOAc to afford the title compound as an off-white solid. ¹H NMR(DMSO-d₆, 400 MHz) δ 8.56 (2H, m), 8.10 (2H, m), 7.84 (1H, d, J=8.0 Hz),7.72 (1H, d, J=8.0 Hz), 7.42 (2H, m), 7.33 (1H, m), 7.22 (1H, d, J=7.6Hz), 4.50 (2H, br s), 4.17 (2H, br s), 2.96 (2H, q, J=7.2 Hz), 2.80 (3H,s), 2.06 (3H, s), 1.27 (3H, t, J=7.2 Hz). LC/MS (Method A): 510 (M+H)⁺.HPLC (Method G) Rt 3.35 min (Purity: 94.9%).

Example 156 In Vitro Assays

Membranes Preparation:

Membranes were prepared from CHO cells expressing S1P1 or S1P3 for usein ligand and 355-GTPγS binding studies. Cells were suspended in 50 mMTRIS, pH 7.4, 2 mM EDTA, 250 mM Sucrose (buffer A) and 1× Completeprotease inhibitor cocktail (Roche), and disrupted at 4° C. by N2decompression using a cell disruption bomb (Parr Instrument). Followingcentrifugation at 1000 RPM for 10 min at 4° C., the supernatant wasdiluted (2×) in buffer A and centrifuged again at 19000 RPM for 75 minat 4° C. The pellet was then suspended in 10 mM HEPES, pH 7.4, 1 mMEDTA, 250 mM Sucrose (Buffer B), and 1× Complete EDTA-free proteaseinhibitor cocktail and homogenized using a potter. Membranes were flashfrozen in liquid N₂ and stored at −80° C.

Receptor Binding Assay:

[33P]sphingosine 1-phosphate (3000 Ci/mmol; American RadiolabeledChemicals, Inc.) was added to test compounds in 20% DMSO by competition.Membranes and WGA SPA beads (GE Healthcare) were added to give a finalvolume of 100 μl in 96-well plates or 50 μl in 384-well plates withassay concentrations of 30 pM or 15 pM [33P]sphingosine 1-phosphate(respectively for S1P1 or S1P3), 50 mM HEPES, pH 7.5, 5 mM MgCl₂, 100 mMNaCl, 0.4% fatty acid-free BSA, 1-5 μg/well of proteins in 96-wellplates vs 0.6-1 μg/well of proteins in 384-well plates and 100 μg/wellof WGA SPA beads in 96-well plates vs 75 μg/well of WGA SPA beads in384-well plates. Binding was performed for 60 min at RT on a shaker andbound radioactivity was measured on a PerkinElmer 1450 MicroBetacounter. Triplicate samples were averaged and normalized as percentageof inhibition relative to total binding (only DMSO in well) andnon-specific binding (1000-fold excess of unlabeled S1P). Binding datawere analyzed using the GraphPad Prism program or Genedata software.

Measurements of 35S-GTPγS Binding:

Membranes (1 to 10 μg protein) prepared as described above, wereincubated in 96-well Scintiplates (PerkinElmer) with test compoundsdiluted in DMSO, in 140 μl of 20 mM HEPES, pH 7.4, 10 mM MgCl2, 2μg/well Saponin, 0.2% fatty acid free BSA (Assay buffer), 125 mM NaCland 1.5 μM GDP. The assay was initiated with the addition of 60 μl of1.5 nM [35S]-GTPγS (1100 Ci/mmol; GE Healthcare) in assay buffer. After60 min incubation at 30° C. on a shaker, plates were centrifuged for 10min at 2000 RPM. Supernatant was discarded and membrane boundradioactivity was measured on a PerkinElmer 1450 MicroBeta counter.Triplicate samples were averaged and expressed as % response relative toS1P activation in absence of compound (n=2).

Cellular functional assays: Internalization of Sphingosine-1-phosphatereceptor 1 (S1P₁) in a human Cell line (U2OS) in a 384-well format usinga Cell Imaging analysis.

Jo, E.; Sanna, M. G.; Gonzalez-Cabrera, P. J.; Thangada, S.; Tigyi, S.;Osborne, D. A.; Hla, T.; Parrill, A. L.; Rosen, H. Chem. Biol. 2005, 12,703

The S1P₁ internalization assay was performed in 384 well plates(Corning® 384 black with clear bottom 3712) using S1P₁-U2OS cells fromBiolmage (C039A), a human epithelial cell line (Human Bone OsteosarcomaEpithelial Cells). These cells expressed the human S1P₁ Receptor fusedto the green fluorescent protein (EGFP). A standard CMV promoter(cytomegalovirus promoter) controls the expression of S1P1-EGFP andcontinuous expression was maintained by addition of geneticin to theculture medium.

S1P₁Receptor desensitization induced the internalization of themembrane-localized S1P₁-EGFP fusion protein to endosomes, which can bemonitored by cell imaging analysis. The cells are plated in low serummedium (Dulbecco's Modified Eagle Medium (DMEM) with Glutamax-1 and highglucose, 1% Penicillin/Streptomycin, 1% Fetal Calf Serum (FCS), 0.5mg/ml Geneticin) overnight.

The next day, S1P₁-U2OS cells are incubated in 20 μl serum free medium(DMEM with Glutamax-1 and high glucose, 0.1% of fatty-acid free BovinSerum Albumin (BSA), 10 mM, N′-2-Hydroxyethylpiperazine-N′-2ethanesulphonic acid (HEPES) 1M) for 2 hours at 37° C./5% CO₂. The cellsare then treated with 4 μl compounds/agonists (6×/3% DMSO) for a totalvolume of 24 μl, and plates are incubated for 1 hour at 37° C./5% CO₂.

S1P₁-U2OS cells are fixed with 25 μl Paraformaldehyde 8% and stainedwith Hoechst 33345 dye (1:1000) for 20 minutes.

They were then washed 3 times with Phosphate Buffered Saline (PBS) andplates are sealed. The internalization of the receptor S1P₁-EGFP ismeasured on Cellomics by calculating the “spot count per object”(“object” corresponds to nuclear and “spot” corresponds to S1P₁-EGFPreceptor). Internalization data were observed thanks to vHCS View andanalyzed using Genedata® software.

The compounds of formula (I) have utility as immunoregulatory agents asdemonstrated by their activity as potent agonists of the S₁P₁ receptor,as measured in the assays described above. EC₅₀ of the compounds offormula (I) and subformulae for S₁P₁ is below 0.1 μM. Preferredcompounds of formula (I) exhibit an EC₅₀ for S1P1 receptor below 0.01μM. More preferred compounds of Formula (I) exhibit EC₅₀ for S₁P₁ below0.001 μM. Compounds of formula (I) exhibit a selectivity for the S₁P₁receptor over the S₁P₃ receptor as measured by the ratio of EC₅₀ for theS₁P₁ receptor to the EC₅₀ for the S₁P₃ receptor as evaluated in the³⁵S-GTP'γS binding assay described above. The ratio of EC50 S₁P₁ to EC50S₁P₃ is more than 20, preferably more than 50, more preferably more than100 and even more preferably more than 1000.

The “potency” or the “activity” of the compounds is determined by theEC50 values as evaluated in the above described 35S-GTPγS binding assay.The lowest EC50 values characterize the most potent or active compounds,according to the present invention.

The following results have been obtained:

S1P1 S1P3 S1P1 S1P3 S1P1 S1P1/G S1P3/G binding binding binding bindinginternal- TPG TPG Ki (M) Ki (M) Ki (M) Ki (M) ization EC50 EC50 (96well- (96 well- (384 well- (384 well- EC50 Nb structure (M) (M) plate)plate) plate) plate) (M) 14

1.24E−09 — 4.47E−10 2.36E−07 1.93E−09 1.68E−07 8.75E−09 49

— — 8.70E−10 1.50E−06 2.35E−09 1.41E−06 1.13E−08 50

— — 8.49E−10 7.71E−07 1.24E−08 1.18E−06 2.53E−08 64

— — 2.96E−10 3.68E−08 — — 8.11E−09 67

— — 2.31E−10 3.69E−08 — — 1.19E−08 68

— — 3.57E−10 1.09E−07 — — 1.12E−08 69

— — 2.21E−10 4.67E−08 1.47E−09 — 8.97E−09 70

— — 6.81E−10 4.43E−07 2.19E−09 4.11E−07 1.30E−08 71

— — 1.78E−10 1.31E−07 — — 1.35E−09 72

— — 2.70E−10 1.88E−07 — — 2.05E−09 81

— — 4.12E−10 8.31E−07 2.74E−09 8.55E−07 1.14E−08 89

— — 3.49E−10 6.29E−07 1.19E−09 5.69E−07 6.53E−09 95

— — 4.26E−10 6.50E−07 3.67E−09 1.03E−06 1.39E−08 97

— — — 1.57E−06 2.08E−09 2.22E−06 1.20E−08 100

— — 1.04E−09 9.43E−06 3.49E−09 9.67E−06 5.61E−08 101

— — — 9.43E−06 5.63E−09 6.59E−06 6.93E−08 103

— — — 4.12E−07 1.45E−09 4.68E−07 1.77E−08 104

— — — — 8.98E−09 2.14E−06 1.50E−08 105

— — — 1.83E−06 3.30E−09 1.88E−06 7.49E−08 109

— — — — 1.50E−09 2.93E−07 1.19E−08 110

— — — — 6.22E−09 5.67E−08 111

— — — — 3.68E−09 4.59E−08 112

— — — 1.49E−07 4.95E−10 1.92E−07 6.26E−09 113

— — — 2.17E−06 2.90E−09 2.70E−06 2.48E−08 114

— — — 1.09E−06 1.91E−09 9.14E−07 1.56E−08 115

— — — 6.68E−07 2.15E−09 2.21E−06 9.03E−09 116

— — — 3.33E−07 1.33E−09 1.31E−06 3.65E−09 117

— — — — 3.24E−09 5.04E−08 123

— — — — 3.46E−09 6.90E−06 3.04E−08 129

— — — — 5.33E−09 9.76E−07 2.33E−08 134

— — — — 4.07E−09 1.80E−05 3.73E−08 135

— — — — 2.63E−09 2.61E−08 136

— — — — 4.27E−09 2.32E−06 4.09E−08 138

— — — — 5.16E−09 4.79E−07 2.61E−08 145

— — — — 2.22E−09 2.85E−07 9.44E−09 148

— — — — 2.38E−09 8.34E−07 2.47E−08 150

— — — — 2.07E−09 2.97E−07 1.80E−08 152

— — — — 3.71E−09 1.25E−05 6.84E−08 153

— — — — 3.32E−09 7.50E−06 5.09E−08 154

— — — — 3.16E−09 1.07E−06 1.73E−08 155

— — — — 3.72E−09 8.10E−06 1.73E−08

Example 157 Animal Models Evaluating the In Vivo Efficacy of S1PAgonists Model of SIP Agonists-Induced Lymphopenia in Mice

Female C57BL/6 mice (Elevage Janvier) (8 week old) receive S1P agonistsby oral route. Blood is sampled in heparinized (100 IU/kg, ip) mice byintracardiac or retroorbital puncture under isoflurane anesthesia 2 to120 hrs after drug treatment. The white blood cells (lymphocytes andneutrophils) are counted using a Beckman/Coulter counter. The quality ofblood sampling is assessed by counting erythocytes and platelets.

Model of MOG-Induced Experimental Autoimmune Encephalomyelytis (EAE) inMice

EAE was induced in 9 weeks old female mice (C57BL/6, Elevage Janvier) byan immunization against MOG. The mice received Pertussis toxin (Alexis,300 ng/mouse in 200 μl of PBS) by ip route and 100 μl of an emulsioncontaining MOG35-55 peptide (NeoMPS, 200 μg/mouse), MycobacteriumTuberculosis (0.25 mg/mouse) in Complete Freund's Adjuvant (DIFCO) bysubcutaneous injection into the back. Two days later an additionalinjection of Pertussis toxin (Alexis, 300 ng/mouse in 200 μl of PBS) wasdone by ip route. After EAE induction, mice were weighed daily and theneurological impairment was quantified using a 15-points clinical scaleassessing the paralysis (tail, hind limbs and fore limbs), theincontinency and the death.

Pharmacokinetics Data:

The pharmacoinetic properties of compound of example 70,N-(2-fluoro-4-{5-[2′-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)-N-methylglycine,are the following:

% of lymphopenia in mouse at 48 h 72 +/− 8 PK parameters fromlymphopenia (mouse) PK-PD (30 mpk) Plasma AUC∞ (h * ng/ml) 103845 Cmax(ng/ml) 8780 Tmax (h) 2 T1/2 (h) 7.5 Cl/F (L/kg/h) 0.3 Brain/plasmaratio: AUCZ (h * ng/ml) 2.4 Lymphe node/plasma ratio: 24 h/48 h 4.4/4.0

Clinical Score

-   -   1—Tail        -   Score=0 A normal mouse holds its tail erect when moving.        -   Score=1 If the extremity of the tail is flaccid with a            tendency to fall.        -   Score=2 If the tail is completely flaccid and drags on the            table.    -   2—Hind limbs        -   Score=0 A normal mouse has an energetic walk and doesn't            drag his paws.        -   Score=1 Either one of the following tests is positive:    -   a—Flip test: while holding the tail between thumb and index        finger, flip the animal on his back and observe the time it        takes to right itself. A healthy mouse will turn itself        immediately. A delay suggests hind-limb weakness.    -   b—Place the mouse on the wire cage top and observe as it crosses        from one side to the other. If one or both limbs frequently slip        between the bars we consider that there is a partial paralysis.        -   Score=2 Both previous tests are positive.        -   Score=3 One or both hind limbs show signs of paralysis but            some movements are preserved; for example: the animal can            grasp and hold on to the underside of the wire cage top for            a short moment before letting go        -   Score=4 When both hind legs are paralyzed and the mouse            drags them when moving.    -   3—Fore limbs:        -   Score=0 A normal mouse uses his front paws actively for            grasping and walking and holds his head erect.        -   Score=1 Walking is possible but difficult due to a weakness            in one or both of the paws, for example, the front paws are            considered weak when the mouse has difficulty grasping the            underside of the wire top cage. Another sign of weakness is            head drooping.        -   Score=2 When one forelimb is paralyzed (impossibility to            grasp and the mouse turns around the paralyzed limb). At            this time the head has also lost much of its muscle tone.        -   Score=3 Mouse cannot move, and food and water are            unattainable.    -   4—Bladder:        -   Score=0 A normal mouse has full control of his bladder.        -   Score=1 A mouse is considered incontinent when his lower            body is soaked with urine.    -   5—Death:        -   Score=15

The final score for each animal is determined by the addition of all theabove-mentioned categories. The maximum score for live animals is 10.

At day 12 (first signs of paralysis) the mice were stratified inexperimental groups (n=10) according to the clinical score and the bodyweight loss. The semi-curative treatment started at day 14.

Example 158 Preparation of a Pharmaceutical Formulation

Formulation 1—Tablets

A compound of formula (I) is admixed as a dry powder with a dry gelatinbinder in an approximate 1:2 weight ratio. A minor amount of magnesiumstearate is added as a lubricant. The mixture is formed into 240-270 mgtablets (80-90 mg of active compound according to the invention pertablet) in a tablet press.

Formulation 2—Capsules

A compound of formula (I) is admixed as a dry powder with a starchdiluent in an approximate 1:1 weight ratio. The mixture is filled into250 mg capsules (125 mg of active compound according to the inventionper capsule).

Formulation 3—Liquid

A compound of formula (I) (1250 mg), sucrose (1.75 g) and xanthan gum (4mg) are blended, passed through a No. 10 mesh U.S. sieve, and then mixedwith a previously prepared solution of microcrystalline cellulose andsodium carboxymethyl cellulose (11:89, 50 mg) in water. Sodium benzoate(10 mg), flavor, and color are diluted with water and added withstirring. Sufficient water is then added to produce a total volume of 5mL.

Formulation 4—Tablets

A compound of formula (I) is admixed as a dry powder with a dry gelatinbinder in an approximate 1:2 weight ratio. A minor amount of magnesiumstearate is added as a lubricant. The mixture is formed into 450-900 mgtablets (150-300 mg of active compound according to the invention) in atablet press.

Formulation 5—Injection

A compound of formula (I) is dissolved in a buffered sterile salineinjectable aqueous medium to a concentration of approximately 5 mg/mL.

The invention claimed is:
 1. An oxadiazole derivative of Formula (AC)

wherein R¹, R² independently from one another denote H, CF₃, OCF₃, CN,NO₂, or OH, S is COOH, Q denotes (CH₂)X(CH₂), X is —NA-, W denotes CH,G₁, G₂ denote H, R^(s) is CH₃ or CH₂CH₃, R^(b) is CF₃, A denotes CH₃,CH(CH₃)₂ or CH₂CH₃, and pharmaceutically acceptable salts andstereoisomers thereof.
 2. The compound according to claim 1, whereinsaid compound is selected from: Example Nb structure 14

81

103

109

113

117

154

155

and pharmaceutically acceptable salts and stereoisomers thereof.
 3. Apharmaceutical composition comprising at least one compound according toclaim 1 and a pharmaceutically acceptable excipient.
 4. Thepharmaceutical composition according to claim 3, said compositioncomprising at least one further active ingredient.
 5. A kit consistingof separate packs of (a) an effective amount of a compound according toclaim 1; and (b) an effective amount of a further medicament activeingredient.
 6. A method of treating an autoimmune disorder orimmunoregulatory abnormality comprising the administration of aneffective amount of a compound according to claim 1 to a subject havingan autoimmune disorder or immunoregulatory abnormality.
 7. The methodaccording to claim 6, wherein said immunoregulatory abnormality orautoimmune disorder is selected from the group consisting of: systemiclupus erythematosis, chronic rheumatoid arthritis, inflammatory boweldisease, multiple sclerosis, amyotrophic lateral sclerosis (ALS),arteriosclerosis, atherosclerosis, scleroderma, and autoimmunehepatitis.